Methods of cancer detection

ABSTRACT

The present invention provides, among other things, methods of identifying cancerous or pre-cancerous tissue including providing a first region of tissue from a subject, calculating a roughness exponent for the first region of tissue, and comparing the roughness exponent of the first region of tissue to 0.5, wherein a difference of less than 0.2 between the roughness exponent of the first region of tissue and 0.5 indicates that the tissue is cancerous or pre-cancerous. Additionally, the present invention provides methods including providing a first view of a region of tissue, providing a second view of a region of tissue, calculating a first fractal dimension for the first view of the region of tissue, and calculating a second fractal dimension for the second view of the region of tissue, wherein if the fractal dimension of at least one of the first fractal dimension and the second fractal dimension is in the fractal zone, the region of tissue is considered cancerous. Also provided are systems for performing these assessments.

CROSS-REFERENCE TO RELATED APPLICATIONS

This patent application is a divisional application of U.S. applicationSer. No. 14/786,366, filed Oct. 22, 2015, now issued U.S. Pat. No.10,467,755, which was a U.S. National Stage Entry of InternationalPatent Application No. PCT/US2014/035153, filed Apr. 23, 2014, whichclaims priority to U.S. Provisional Application No. 61/815,209, filedApr. 23, 2013, the contents of each of which are hereby incorporated byreference in their entirety for all purposes herein.

BACKGROUND

The human body is composed of about ten trillion cells, and throughout alifetime cells go through many damaging processes that concern theirgenetic programming. However, not all of these mutations result incancer. As it turns out, many individuals actually have occult tumorsthroughout their bodies that are only discovered through microscopicinvestigation during autopsy. Therefore, there must be some mechanismthat prohibits the growth and development of tumors.

Recent research has pointed to the microenvironment of potential tumorsto help suppress malignant phenotype and instruct otherwise malignantcells to participate in normal development. The microenvironment mayactually provide tumor suppressive signals as long as the tissuearchitecture is controlled, but as the structure of healthy tissue islost, the tissue has the potential to become a tumor promoter. Thus,considering the tissue surrounding the tumor, along with the lesion, maybe important in identifying potential cancerous tissue.

SUMMARY OF THE INVENTION

The present invention is based, in part, on the realization thatdisorder of the microenvironment surrounding a portion of tissue in asubject may be indicative of the presence of, or pre-disposition towarddeveloping, cancer in that tissue. Among other things, the presentinvention provides, for the first time, methods for analyzing the amountand distribution of disorder across tissue in order to predict thepresence of, or propensity of tissue to develop, cancer. In someembodiments, provided methods include analysis of images of targettissue, as opposed to analysis of the tissue itself In some embodiments,provided methods include analysis of tissue or images of tissue adjacentto a target tissue/tissue region of interest, as opposed to the targettissue itself.

The present invention provides, among other things, methods ofidentifying cancerous or pre-cancerous tissue including providing afirst region of tissue from a subject, calculating a roughness exponent(e.g., a Hurst exponent) for the first region of tissue, and comparingthe roughness exponent of the first region of tissue to 0.5, wherein adifference of less than 0.2 between the roughness exponent of the firstregion of tissue and 0.5 indicates that the tissue is cancerous orpre-cancerous. In some embodiments, the providing, calculating, andcomparing steps are each performed a plurality of times. In someembodiments, the plurality of times is at least 10, at least 100, or atleast 1,000. In some embodiments, performing the providing, calculatingand comparing steps a plurality of times allows for a higher resolutioncharacterization of the morphology of the tissue surrounding a region oftissue suspected to be cancerous or pre-cancerous.

In some embodiments, the difference between the roughness exponentvalues and 0.5 may be less than 0.2. In some embodiments, the differencebetween the roughness exponent of the first region of tissue and 0.5 isless than or equal to 0.15. In some embodiments, the difference betweenthe roughness exponent of the first region of tissue and 0.5 is lessthan or equal to 0.1. In some embodiments, the difference between theroughness exponent of the first region of tissue and 0.5 is less than orequal to 0.05.

In some embodiments, a roughness exponent is calculated from one or moreimages of a target tissue. In some embodiments, a roughness exponent iscalculated from two or more images of a first region of tissue. In someembodiments, each of the two or more images are taken from differentangles and/or points of view. In some embodiments, a roughness exponentis calculated for two or more regions of tissue. In some embodiments,the two or more regions of tissue are from a single organ. In someembodiments, the two or more regions of tissue are from differentorgans.

A roughness exponent may be generated via any appropriate multi-scaleanalytical method. In some embodiments, the roughness exponent iscalculated using one or more multi-scale methods selected from awavelet-transform modulus maxima, a wavelet leader, detrendedfluctuation, and Fourier analysis. A roughness exponent may also becalculated using any mathematically related and/or similar multi-scaledensity fluctuation assessment method yielding an exponent or spectrumof exponents that is similar and/or can complement the use of the Hurstexponent. In some embodiments, a roughness exponent is calculated fromtwo-dimensional data. In some embodiments, a roughness exponent iscalculated from three dimensional data (e.g., 3D data cubes).

In some embodiments, the present invention additionally provides methodsincluding providing a first view of a region of tissue; providing asecond view of the region of tissue; calculating a first fractaldimension for the first view of the region of tissue; and calculating asecond fractal dimension for the second view of the region of tissue;wherein if the fractal dimension of at least one of the first fractaldimension and the second fractal dimension is in the fractal zone, theregion of tissue is considered cancerous. In some embodiments, providedmethods further comprise treating the region of tissue if it iscancerous.

In some embodiments, the fractal zone is defined as a polygon consistingof a central square and a first, second, third, and fourth extendingtriangular region as plotted on a graph of the fractal dimension of thefirst view of the region of tissue by the fractal dimension of thesecond view of the region of tissue.

In some embodiments, the central square and first, second, third andfourth extending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view:

central square: (1.2, 1.2), (1.2, 1.8), (1.8, 1.2)(1.8, 1.8);

first extending triangular region: (0.5, 1.5), (1.2, 1.2), (1.2, 1.8);

second extending triangular region: (1.5, 0.5), (1.2, 1.2), (1.8, 1.2);

third extending triangular region: (1.5, 2.3), (1.2, 1.8), (1.8, 1.8);and

fourth extending triangular region: (2.3, 1.5), (1.8, 1.2), (1.8, 1.8).

In some embodiments, the central square and first, second, third andfourth extending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view:

central square: (1.1, 1.1), (1.1, 1.9), (1.9, 1.1)(1.9, 1.9);

first extending triangular region: (0.5, 1.5), (1.1, 1.1), (1.1, 1.9);

second extending triangular region: (1.5, 0.5), (1.1, 1.1), (1.9, 1.1);

third extending triangular region: (1.5, 2.3), (1.1, 1.9), (1.9, 1.9);and

fourth extending triangular region: (2.3, 1.5), (1.9, 1.1), (1.9, 1.9).

In some embodiments, the central square and first, second, third andfourth extending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view:

central square: (1.3, 1.3), (1.3, 1.7), (1.7, 1.3)(1.7, 1.7);

first extending triangular region: (0.5, 1.5), (1.3, 1.3), (1.3, 1.7);

second extending triangular region: (1.5, 0.5), (1.3, 1.3), (1.7, 1.3);

third extending triangular region: (1.5, 2.3), (1.3, 1.7), (1.7, 1.7);and

fourth extending triangular region: (2.3, 1.5), (1.7, 1.3), (1.7, 1.7).

In some embodiments, provided methods further include steps of providinga third view of the region of tissue; and calculating a third fractaldimension for the third view of the region of tissue; wherein if thefractal dimension of at least one of the first fractal dimension, thesecond fractal dimension, and the third fractal dimension is in thefractal zone, the region of tissue is considered cancerous. In someembodiments, a fractal dimension is calculated from two-dimensionaldata. In some embodiments, a fractal dimension is calculated from threedimensional data (e.g., 3D data cubes).

The methods provided by the present invention may be performed on anytissue. In some embodiments, the tissue is selected from breast tissue,brain tissue, lung tissue, kidney tissue, liver tissue, uterine tissue,dermal tissue, and pancreatic tissue.

As used in this application, the terms “about” and “approximately” areused as equivalents. Any numerals used in this application with orwithout about/approximately are meant to cover any normal fluctuationsappreciated by one of ordinary skill in the relevant art.

Other features, objects, and advantages of the present invention areapparent in the detailed description that follows. It should beunderstood, however, that the detailed description, while indicatingembodiments of the present invention, is given by way of illustrationonly, not limitation. Various changes and modifications within the scopeof the invention will become apparent to those skilled in the art fromthe detailed description.

Definitions

In order for the present invention to be more readily understood,certain terms are first defined below. Additional definitions for thefollowing terms and other terms are set forth throughout thespecification.

Animal: As used herein, the term “animal” refers to any member of theanimal kingdom. In some embodiments, “animal” refers to humans, at anystage of development. In some embodiments, “animal” refers to non-humananimals, at any stage of development. In certain embodiments, thenon-human animal is a mammal (e.g., a rodent, a mouse, a rat, a rabbit,a monkey, a dog, a cat, a sheep, cattle, a primate, and/or a pig). Insome embodiments, animals include, but are not limited to, mammals,birds, reptiles, amphibians, fish, insects, and/or worms. In someembodiments, an animal may be a transgenic animal,genetically-engineered animal, and/or a clone.

Approximately or about: As used herein, the term “approximately” or“about,” as applied to one or more values of interest, refers to a valuethat is similar to a stated reference value. In certain embodiments, theterm “approximately” or “about” refers to a range of values that fallwithin 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%,8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greaterthan or less than) of the stated reference value unless otherwise statedor otherwise evident from the context (except where such number wouldexceed 100% of a possible value).

Cancer: As used herein, the term “cancer” refers to a group of diseases,all involving unregulated cell growth. Exemplary cancers include,without limitation: Acute lymphoblastic leukemia, Acute myeloidleukemia, Adrenocortical carcinoma; AIDS-related cancers; AIDS-relatedlymphoma; Anal cancer; Appendix cancer; Astrocytoma,childhood cerebellaror cerebral; Basal cell carcinoma; Bile duct cancer, extrahepatic;Bladder cancer; Bone cancer, Osteosarcoma/Malignant fibroushistiocytoma; Brainstem glioma; Brain tumor; Brain tumor, cerebellarastrocytoma; Brain tumor, cerebral astrocytoma/malignant glioma; Braintumor, ependymoma; Brain tumor, medulloblastoma; Brain tumor,supratentorial primitive neuroectodermal tumors; Brain tumor, visualpathway and hypothalamic glioma; Breast cancer; Bronchialadenomas/carcinoids; Burkitt lymphoma; Carcinoid tumor, childhood;Carcinoid tumor, gastrointestinal; Carcinoma of unknown primary; Centralnervous system lymphoma, primary; Cerebellar astrocytoma, childhood;Cerebral astrocytoma/Malignant glioma, childhood; Cervical cancer;Childhood cancers; Chronic lymphocytic leukemia; Chronic myelogenousleukemia; Chronic myeloproliferative disorders; Colon Cancer; CutaneousT-cell lymphoma; Desmoplastic small round cell tumor; Endometrialcancer; Ependymoma; Esophageal cancer; Ewing's sarcoma in the Ewingfamily of tumors; Extracranial germ cell tumor, Childhood; ExtragonadalGerm cell tumor; Extrahepatic bile duct cancer; Eye Cancer, Intraocularmelanoma; Eye Cancer, Retinoblastoma; Gallbladder cancer; Gastric(Stomach) cancer; Gastrointestinal Carcinoid Tumor; Gastrointestinalstromal tumor (GIST); Germ cell tumor: extracranial, extragonadal, orovarian; Gestational trophoblastic tumor; Glioma of the brain stem;Glioma, Childhood Cerebral Astrocytoma; Glioma, Childhood Visual Pathwayand Hypothalamic; Gastric carcinoid; Hairy cell leukemia; Head and neckcancer; Heart cancer; Hepatocellular (liver) cancer; Hodgkin lymphoma;Hypopharyngeal cancer; Hypothalamic and visual pathway glioma,childhood; Intraocular Melanoma; Islet Cell Carcinoma (EndocrinePancreas); Kaposi sarcoma; Kidney cancer (renal cell cancer); LaryngealCancer; Leukemias; Leukemia, acute lymphoblastic (also called acutelymphocytic leukemia); Leukemia, acute myeloid (also called acutemyelogenous leukemia); Leukemia, chronic lymphocytic (also calledchronic lymphocytic leukemia); Leukemia, chronic myelogenous (alsocalled chronic myeloid leukemia); Leukemia, hairy cell; Lip and OralCavity Cancer; Liposarcoma; Liver Cancer (Primary); Lung Cancer,Non-Small Cell; Lung Cancer Small Cell Lymphomas; Lymphoma, Burkitt;Lymphoma, cutaneous T-Cell; Lymphoma, Hodgkin; Lymphoma, Primary CentralNervous System; Macroglobulinemia, Waldenström; Malignant FibrousHistiocytoma of Bone/Osteosarcoma; Medulloblastoma, Childhood; Melanoma;Melanoma, Intraocular (Eye); Merkel Cell Carcinoma; Mesothelioma, AdultMalignant; Mesothelioma, Childhood; Metastatic Squamous Neck Cancer withOccult Primary; Mouth Cancer; Multiple Endocrine Neoplasia Syndrome,Childhood; Multiple Myeloma/Plasma Cell Neoplasm; Mycosis Fungoides;Myelodysplastic Syndromes; Myelodysplastic/Myeloproliferative Diseases;Myelogenous Leukemia, Chronic; Mycloid Leukemia, Adult Acute; MycloidLeukemia, Childhood Acute; Myeloma, Multiple (Cancer of theBone-Marrow); Myeloproliferative Disorders, Chronic; Nasal cavity andparanasal sinus cancer; Nasopharyngeal carcinoma; Neuroblastoma;Non-Hodgkin lymphoma; Non-small cell lung cancer; Oral Cancer;Oropharyngeal cancer; Osteosarcoma/malignant fibrous histiocytoma ofbone; Ovarian cancer; Ovarian epithelial cancer (Surfaceepithelial-stromal tumor); Ovarian germ cell tumor; Ovarian lowmalignant potential tumor; Pancreatic cancer; Pancreatic cancer, isletcell; Paranasal sinus and nasal cavity cancer; Parathyroid cancer;Penile cancer; Pharyngeal cancer; Pheochromocytoma; Pineal astrocytoma;Pineal germinoma; Pineoblastoma and supratentorial primitiveneuroectodermal tumors, childhood; Pituitary adenoma; Plasma cellneoplasia/Multiple myeloma; Pleuropulmonary blastoma; Primary centralnervous system lymphoma; Prostate cancer; Rectal cancer; Renal cellcarcinoma (kidney cancer); Renal pelvis and ureter, transitional cellcancer; Retinoblastoma; Rhabdomyosarcoma, childhood; Salivary glandcancer; Sarcoma, Ewing family of tumors; Sarcoma, Kaposi; Sarcoma, softtissue; Sarcoma, uterine; Sézary syndrome; Skin cancer (nonmelanoma);Skin carcinoma, Merkel cell; Small intestine cancer; Soft tissuesarcoma; Squamous cell carcinoma; Squamous neck cancer with occultprimary, metastatic; Stomach cancer; Supratentorial primitiveneuroectodermal tumor, childhood; T-Cell lymphoma, cutaneous; Testicularcancer; Throat cancer; Thymoma, childhood; Thymoma and Thymic carcinoma;Thyroid cancer; Thyroid cancer, childhood; Transitional cell cancer ofthe renal pelvis and ureter; Trophoblastic tumor, gestational; Unknownprimary site, carcinoma of, adult; Unknown primary site, cancer of,childhood; Ureter and renal pelvis, transitional cell cancer; Urethralcancer; Uterine cancer, endometrial; Uterine sarcoma; Vaginal cancer;Visual pathway and hypothalamic glioma, childhood; Vulvar cancer; andWilms tumor (kidney cancer), childhood.

Pre-Cancer: As used herein the term “pre-cancer” or “pre-cancerous” isused to refer to tissue in or from a subject that is not yet cancerous,but has a higher chance of becoming cancerous that normal tissue.

Risk: As will be understood from context, a “risk” of a disease,disorder, and/or condition comprises a likelihood that a particularindividual will develop a disease, disorder, and/or condition (e.g., acancer). In some embodiments, risk is expressed as a percentage. In someembodiments, risk is from 0,1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40,50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed asa risk relative to a risk associated with a reference sample or group ofreference samples. In some embodiments, a reference sample or group ofreference samples have a known risk of a disease, disorder, conditionand/or event (e.g., a muscular dystrophy). In some embodiments areference sample or group of reference samples are from individualscomparable to a particular individual. In some embodiments, relativerisk is 0,1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.

Subject: As used herein, the term “subject” refers to a human or anynon-human animal (e.g., mouse, rat, rabbit, dog, cat, cattle, swine,sheep, horse or primate). A human includes pre and post natal forms. Inmany embodiments, a subject is a human being. A subject can be apatient, which refers to a human presenting to a medical provider fordiagnosis or treatment of a disease. The term “subject” is used hereininterchangeably with “individual” or “patient.” A subject can beafflicted with or is susceptible to a disease or disorder but may or maynot display symptoms of the disease or disorder.

Substantially: As used herein, the term “substantially” refers to thequalitative condition of exhibiting total or near-total extent or degreeof a characteristic or property of interest. One of ordinary skill inthe biological arts will understand that biological and chemicalphenomena rarely, if ever, go to completion and/or proceed tocompleteness or achieve or avoid an absolute result. The term“substantially” is therefore used herein to capture the potential lackof completeness inherent in many biological and chemical phenomena.

BRIEF DESCRIPTION OF THE DRAWING

FIG. 1 shows a representative schematic of imaging breast tissue frommediolateral oblique (MLO) and cranio-caudal (CC) angles.

FIG. 2 depicts a graph of the minimum fractal dimension by maximumfractal dimension, with tissue classified according to known methods asbenign in blue, and tissue classified according to known methods asmalignant tissue, in red.

FIG. 3 shows exemplary results of a cluster analysis of scatter plotsobtained from graphing minimum fractal dimension by maximum fractaldimension of (a) benign tissue, and (b) malignant tissue.

FIG. 4 shows images of breast tissue classified as (a) benign and (b)malignant as classified by provided methods. Note the presence of amajority of disrupted tissue in the image of malignant tissue(represented by yellow circles) and the lack of disrupted tissue in thebenign tissue (denoted by a lack of yellow circles).

FIG. 5 shows a boxplot of the distribution of AH for both benign andmalignant (cancer) tissues.

FIG. 6 shows exemplary photographs of breast tissue classified accordingto provided methods. Specifically, the left panel shows control breasttissue made up of healthy fatty tissue (H<0.45); the center panel showsa breast with a benign lesion lying at the interface between fatty anddense tissue, showing a few surrounding sub-regions of disrupted tissue(yellow, 0.45≤H≤0.55); and the right panel shows a breast with amalignant lesion with several disrupted tissue neighboring regions.

FIG. 7A shows a sample simulated fractional Brownian motion imageB_(H-0.5)(x); 7B shows the gradient of the image in 7A is obtained asthe modulus of the wavelet transformed using equation 2.10 below; 7Cshows maxima chains in blue defined as positions where the WT modulus islocally maximal (i.e. the WTMM) and along these chains in 7C furtherlocal maxima are found in red (i.e. the WTMMM); this is then repeated asseveral different scales, three of which are shown in 7D, 7E, and 7F;the WTMMM were then connected vertically through scales to define the WTskeleton shown in 7G, with the gray-scale coding going from black(minimum) to white (maximum).

FIG. 8A shows an original image obtained from the DDSM database, and 8Bshows a zoomed in image with a suspicious region encircled by aradiologist; by selecting appropriate values for the slope of h of theWT modulus as a function of scale in a logarithmic representation, andthe log of the pre-factor, log(k) in 8D, the WTMMM (blue) from thetissue background in 8C are distinguished from the WTMMM (red) thatbelong to the MC in 8E; from that point the WT skeleton can becalculated from the WTMMM that belong to the lesion from those thatbelong to the background tissue; the corresponding WTMM chains at thesmallest scale are shown in 8F and 8G for the background and lesion,respectively.

FIG. 9A-G shows the same analysis as in FIG. 8A-G, only on a differentcase.

FIG. 10 shows the frequency distributions of fractal dimensions Dcalculated for benign cranio-caudal (CC) and mediolateral oblique (MLO)views (top panels) as compared to the fractal dimensions D calculatedfor cancer CC and MLO views (bottom panels); it is of note that thedistributions are drastically different between benign and cancersamples.

FIG. 11 shows a fractal dimension plot of several samples; specifically,each case is represented by a single dot and is plotted with the fractaldimension obtained from the mediolateral oblique (MLO) view as afunction of the fractal dimension obtained from the cranio-caudal (CC)view. A polygonal region is outlined, the inside of which is defined asthe “fractal zone” while the outside is defined as the “Euclidean zone”.The dots represent malignant (dark red) and benign (light green) cases.

FIG. 12 shows exemplary random point distribution models to describe howdifferent kinds of objects can have only a limited number of possiblefractal dimension as a function of the projection angle; the left panel(12A) shows the possible fractal dimensions of a line; the center panel(12B) shows the possible fractal dimensions of a surface, and the rightpanel (12C) shows the possible fractal dimensions of a fractal cluster(here a simulated diffusion-limited aggregate).

FIG. 13 shows a block diagram of an exemplary cloud computingenvironment suitable for use with provided methods.

FIG. 14 is a block diagram of an exemplary computing device and a mobilecomputing device suitable for use with provided methods.

DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS

The present invention provides, among other things, methods and systemsfor identifying cancerous or pre-cancerous tissue in a subject. Thepresent invention is based, in part, on the surprising discovery thatanalysis of tissue surrounding a region of interest rather than tissuefrom the region of interest itself, may be used to determine if aparticular region of tissue is cancerous or pre-cancerous. In someembodiments, the present invention provides, among other things, methodsof identifying cancerous or pre-cancerous tissue including providing afirst region of tissue from a subject, calculating a roughness exponentfor the first region of tissue, and comparing the roughness exponent ofthe first region of tissue to 0.5, wherein a difference of less than 0.2(e.g., less than 0.1, less than 0.05) between the roughness exponent ofthe first region of tissue and 0.5 indicates that the tissue iscancerous or pre-cancerous. In some embodiments, provided methodsfurther comprise treating the tissue adjacent to the first region oftissue if it is cancerous or pre-cancerous.

In some embodiments, the present invention also provides methodsincluding providing a first view of a region of tissue, providing asecond view of a region of tissue, calculating a first fractal dimensionfor the first view of the region of tissue, and calculating a secondfractal dimension for the second view of the region of tissue, whereinif the fractal dimension of at least one of the first fractal dimensionand the second fractal dimension is in the fractal zone, the region oftissue is considered cancerous.

In some embodiments, it may be advantageous to perform the providing,calculating, and/or comparing steps of provided methods multiple times.In some embodiments, it is contemplated that performing the providing,calculating and comparing steps multiple times will allow for superiorcharacterization of the tissue or the tissue surrounding a tissue regionof interest as compared to performing each step a single time accordingto various embodiments. In some embodiments, the providing, calculating,and comparing steps are each performed a plurality of times. In someembodiments, the plurality of times is at least 10, at least 20, atleast 50, at least 100, at least 300, at least 500, or at least 1,000times. In some embodiments, performing the providing, calculating, andcomparing steps multiple times occurs for different regions of the sameorgan or tissue. In some embodiments, performing the providing,calculating, and comparing steps multiple times occurs for substantiallythe same region(s) of the same organ or tissue. In some embodiments,performing the providing, calculating, and comparing steps multipletimes occurs for different organs or tissues.

Prior Detection Methods

Breast cancer is the most common cancer worldwide according to the WorldHealth Organization (WHO) and the second leading cause of cancer relateddeath among women in the United States. Despite the recent advances inthe medical field, the breast cancer rate has continued to increase overthe last 30 years. Cancer is easiest to treat when it is found in theearly stages of development, making it critical for women to haveregular screenings as recommended by the American Cancer Society (ACS).Mammograms are currently one of the most accepted screening processesand have been proven to be successful in detecting microcalcifications(MC), which are small deposits of calcium in breast tissue (about 200microns in size), and can be an indicator of the early development ofbreast cancer.

By identifying early breast cancer, the survival rate for the patientincreases. Thus, early detection is key for the patient. However,detecting tumors in the early stages of development may not always beeasy due to complex tissue composition and the small size of the tumor.Therefore, it's important for radiologists to identify any asymmetricchanges in tissue composition in the breast, as it could be a sign of apotential tumor environment. Doctors rely heavily on the use ofmammography, especially for screening of older women, making it a verywidely accepted form of breast cancer detection.

Although mammograms are currently the most effective way of detectingbreast cancer, it remains difficult to interpret mammograms due to hightissue variability and 3D to 2D projection effects. Current practice isto have two expert radiologists read the mammograms to reduceinterpretation errors. However, it is not always possible to have accessto two radiologist's interpretation due the size of the hospital or thecost. Computer Aided Diagnosis (CAD) systems were developed to assistdoctors in analyzing medical images such as mammograms and since theapproval of CAD by the Food and Drug Administration (FDA) in 1998 therehas been much attention to developing such software. However, many ofthe current CAD methods rely on tissue homogeneity characterized by theHurst exponent, meaning the tissue is assumed to be monofractal anduncorrelated. Healthy fatty and dense tissue, shown by dark and lightintensities respectively on mammograms, demonstrate how tissue may notbe statistically homogeneous and there are many fluctuations ofintensity throughout the image. By neglecting to accurately characterizethe environment of possible breast lesions, important information islost. Therefore, many CAD methods are not offering the expectedperformance, which leads to increase recall rates and can causefalse-positives on up to 70% of normal cases, resulting in an increaseof unnecessary stress on women.

A key insight to developing a successful CAD is given by recent researchwhich has shown the importance the microenvironment can have on thegrowth and development of tumors. The environment of cancerous lesionshas shown to be able to create a niche around the stem cells, favoringthe survival of cancerous stem cells and protecting cells from anytreatment or therapy. Thus, differences between normal and cancer stemcells and their interactions with the neighboring tissue may exist andbe able to be detected by a CAD at a much larger scale. Therefore, it iscritical for the neighboring tissue to be carefully examined and takeninto consideration when using or developing a CAD to evaluatemammograms, something not done prior to the present invention.

One limitation of existing CAD is the inability of the software to ratethe significance of the findings, similar to the BreastImaging-Reporting and Data-System (BI-RADS) assessment score given bythe radiologist at the time of the mammogram interpretation. Asdescribed herein, the two scores provided by the 2D Wavelet-TransformModulus Maxima (WTMM) method, i.e. the fractal dimension of the MC andthe roughness of the tissue surrounding the breast lesion given by theHurst exponent, have known physical properties and can provide aninsight to the invasiveness and severity of the possible lesion. Indeed,one of the several advantages provided by the present invention is therecognition that by examining not only the tumor, but also themicroenvironment, important aspects of benign and malignant processescan be evaluated and possibly lead to detection of early cancer or evenpre-cancerous tissue.

Characterization of Dense and Fatty Tissues

An important part of interpreting mammograms relies on the ability ofradiologists to identify the composition of tissue determined by denseand fatty components. Genetics play a role in determining breast densityand radiologists can detect dense tissue as light intensities on anx-ray and fatty tissue as dark intensities. Density decreases with ageas a normal process. To help provide radiologists with a uniform scoringsystem, the American College of Radiology developed an index which ranksdensity from 1 to 4, ranging from fatty to dense as shown in Table 1.

TABLE 1 Breast Density Score Score Definition 1 Fatty Tissue 2 ScatteredFibroglandular 3 Heterogeneously Dense 4 Dense Tissue

By taking into account the tissue composition of the breast,radiologists can more accurately identify suspicious regions. Sincetissue composition should be symmetric between both breasts, differingdensities maybe a potential sight for a tumor, causing radiologists topay close attention to these differing areas.

There have been many statistical studies devoted to mammography analysisby using fractal techniques. One such study was conducted by P. Kestenerand colleagues using the 2D WTMM method to analyze normal mammaryparenchyma. The goal of the Kestener et al. study was to accuratelyclassify mammographic tissue as dense or fatty. There, the images wereobtained from the Digital Database for Screening Mammography (DDSM) atthe University of South Florida. The databank contains over 2,500studies made up of normal, benign and malignant mammograms allcategorized by an expert radiologist. Each study has two images of eachbreast, consisting of a mediolateral oblique (MLO) view andcranio-caudal (CC) view with any suspicious region circled by aradiologist. The suspicious region could contain a mass and/ormicrocalcifications (MC), but only the cases that were classified asnormal, i.e. no suspicious area was identified, were looked at in theKestener et al. study.

After Kestener analyzed 10 images, 5 fatty breasts and 5 dense breasts,from the DDSM website using the 2D WTMM method, their analysischaracterized fatty tissue as H=0.25±0.05 and dense tissue asH=0.65±0.05.

Both fatty and dense tissue display monofractal scaling behavior, withfatty tissue, H=[0.20−0.35] being the signature of anti-persistentroughness fluctuations and dense tissue, H=[0.55−0.75], the signature ofpersistent long-range correlations. Note that no tissue classificationindex exists in H=(0.35−0.65) and healthy breast tissue is composed ofonly fatty or dense tissue.

Application of Wavelet-Transform Modulus Maxima to Subject Tissue

Various embodiments of the present invention apply wavelet-transformmodulus maxima (WTMM) to one or more images of a tissue in order tocharacterize one or more attributes of the tissue and/or tissue adjacentto the tissue (i.e., the tissue's microenvironment). The presentinvention evidences the potential of the two dimensional WTMM method tobecome a powerful tool in interpreting mammograms. The WTMM method hasproven to be successful in several fields of applied science, includinggeology, astrophysics, cellular biology and orthopedic medicine. Themethod was originally developed as a multifractal formalism to analyzehighly complex 1D signals, 2D images, and 3D images. As described indetail herein, the wavelet transform (WT) acts as a mathematicalmicroscope to characterize spatial image information over a continuousrange of size scales. It is the gradient vector of the image smoothed bydilated versions of a Gaussian filter. At each size scale, the wavelettransform modulus maxima (WTMM) are defined by the positions where themodulus of the WT is locally maximal. These WTMM are automaticallyorganized as maxima chains at the considered scale. Along each of thesechains, further local maxima are found, the WTMM maxima (WTMMM). Thisprocess is repeated for all size scales and the WTMMM from each scaleare then linked to form the WT skeleton.

The ability to consider vertical lines in the WT skeleton individuallyis significant, since it allows one to objectively discriminate betweenlines pointing to the tissue background from those pointing to thelesion by considering how the WT modulus varies as a function of thescale parameter along each line. One can then calculate the so-calledsingularity spectrum separately for each subset, which then allowsconsideration of the roughness exponent H, characterizing the tissuebackground, and the fractal dimension D of the lesion, characterizingthe architecture of the lesion, and their microenvironment.

In some embodiments, provided methods make use of an adaptation of a twodimensional WTMM method, specifically, the continuous wavelet transformmethod, as a mathematical microscope used to characterize the fractalgeometry of clusters of microcalcifications (MC) in human tissue (e.g.,breast tissue) and/or to determine the roughness of the backgroundtissue (i.e. tissue adjacent to the tissue of interest) seen inmammograms. The WTMM method yields the so-called singularity spectrum,D(h), i.e. the fractal dimension D, of points having a Holder exponentof h. The MC are seen as Dirac singularities by the WTMM method,therefore having Holder exponent value of h=−1. This allows the MC withh˜−1 to be abstracted from the background tissue which has h˜0.30 forfatty breast tissue and h˜0.65 for dense tissue. Thus, the WTMM methodis used to perform a segmentation of tissues, in some embodiments, thebreast tumor tissue, based on the strength of the singularitiescomposing the mammogram images, and to simultaneously quantify theirfractal dimension.

Comparing the results of provided methods applied on several hundredimages from a digital databank of mammograms with known radiologistdiagnostics, the fractal dimensions of benign and malignant breastlesions are significantly different, with benign having an integerdimension corresponding to a non-invasive Euclidean object and cancerhaving a non-integer dimension, representing an invasive structure. Inaddition, the microenvironments characterized by the roughness of thetissue in which the lesions are embedded (i.e., adjacent to) aredifferent for benign and malignant tumors, and provides an insight intothe onset and development of breast cancer.

One score provided by the two dimensional WTMM method is the roughnessof the tissue surrounding the breast lesion, given by the Hurstexponent, which has known physical properties and can provide an insightto the invasiveness and severity of the possible lesion. By examiningnot only the tumor but also the microenvironment, important aspects ofbenign and malignant processes can be evaluated and possibly lead toearly detection of cancer or even pre-cancerous tissue. In variousembodiments, other multi-scale methods may also be used as analternative to WTMM including, but not limited to, wavelets leaders,detrended fluctuation analysis, Fourier transform methodologies, and/orany mathematically related and/or similar multi-scale densityfluctuation assessment methods yielding an exponent or spectrum ofexponents that is similar and/or can complement the use of the Hurstexponent.

The Hurst exponent (or equivalent roughness exponents obtained bysimilar multi-scale analytical methods) represents, in part, thephysical status of the microenvironment of the tissue (e.g., tissuedensity as determined from roughness fluctuations) and the level ofcorrelation in that physical status across adjacent tissue. In general,if H<0.5, then the roughness fluctuation are considered anti-correlated,while H>0.5 means that the roughness fluctuations are positivelycorrelated. In either case, the physical system of the analyzed tissueis considered to have some form of spatial memory. An H=0.5 means thatthe roughness fluctuations are un correlated and are thus considered tohave no spatial memory.

The present invention encompasses the recognition that an H=0.5, or an Hvalue within some range surrounding 0.5, in some embodiments betweenabout 0.3-0.65, between about 0.35-0.6, between about 0.4-0.6, betweenabout 0.4-0.55, or between about 0.45-0.55, indicates a region of tissuethat is potentially cancerous or pre-cancerous. As described above,around H=0.5 the system has a reduced or loss of correlation betweenroughness fluctuations, indicating a breakdown in order in the system.It is the recognition that this disorder likely indicates a cancerous orpre-cancerous state that is an important aspect of the methods andsystems of the present invention.

The present invention recognizes that the results obtained by using theWTMM method on screening mammograms could not only be used as a possiblecomputer-aided detection (CAD) method, but also as a method to furtherstudy the biophysics of tumor onset and progression. As described in theExamples below, after all images were analyzed using provided methods, astatistical analysis was performed on all data, which provided us withinformation on the critical differences between the organization,behavior, and biological and physical processes of both benign andmalignant tumors.

Exemplary Mechanics of Characterizing the Local Regularity of RoughSurfaces with the Wavelet Transform Modulus Maxima Method

Most of the fractal methods used to analyze mammograms rely on theestimate of the fractal dimension which is related to the Hurst exponentH which statistically characterizes the global roughness of themammogram landscape. The two dimensional WTMM method accounts forpossible fluctuations of the local regularity of a rough surface asdefined by the Holder exponent, h of the function ƒ whose graph definesthe rough surface under study. The 2D WTMM method provides a way toestimate the D(h) singularity spectrum which provides us with theHausdorff dimension of the set of points x where the local roughness ofthe exponent h(x) is h.

This methodology has the ability to be applied to a variety of roughsurfaces, such as mammogram landscapes. We will use the term “roughsurface” for an irregular surface. This means the surface can beaccurately described by a single-valued, self-affine function satisfyingthe following:∀|x ₀=(x ₀ , y ₀) ∈

² , ∀x=(x, y) ∈

² in the neighborhood of xo, ∃H∈

suchthat, for any λ>0:ƒ(x ₀ +λx, y ₀ +λ ^(a) y)−ƒ(x ₀ , y ₀)≅λ^(H)[ƒ(x ₀ +x, y ₀ +y)−ƒ(x ₀ , y₀)]   (2.1)

In various embodiments, the Hurst exponent is used to characterize theglobal roughness of the function under investigation. If H<1, then thefunction, f, is nowhere differentiable and the smaller the value for H,the more singular and irregular f. Several methods have been used toestimate the Hurst exponent of self-affine functions, but one has to becareful due to the fact that some functions, such as fractal functions,generally display multi-affine properties in the sense that theirroughness fluctuates from point to point. To describe these multifractalfunctions, one needs to change the definition of the Hurst exponent of ƒto become a local value, h(x₀). Here, h is called the Holder exponentand it provides the strength of the singularity of ƒ at x₀. A rigorousdefinition of the Holder exponent is given by the largest exponent h(x₀)such that there exists a polynomial of degree n<h(x₀) and a constantC>0, such that for any point x in the neighborhood of x₀ one has:|ƒ(x)−P _(n)(x−x ₀)|≤C|x−x ₀|^(h(x) ⁰ ⁾   (2.2)

If ƒ is n times continuously differentiable at the point x₀, then onecan use the order n Taylor series of ƒ at x₀ for the polynomial Pn(x−x0)and prove h(x₀)>n. Thus h(x0) measures how irregular the function f is.

Multi-Scale Edge Detection

The edges of the different structures that appear in an image are oftenthe most important features for pattern recognition. Therefore, manycomputer tools for edge detection look for points where the gradient ofthe image intensity has a modulus locally maximum in its direction. Withan appropriate choice of analyzing wavelets, one can redefine theCanny's multi-scale edge detection in terms of a 2D wavelet transform.

In several embodiments, use of a multi-scale edge detection methodologyis desirable to assist in determining the borders of cancerous orpre-cancerous tissue. Below is a brief discussion of exemplarymulti-edge detection techniques that may be used in accordance with thepresent invention.

First, define two wavelets that are the partial derivatives with respectto x and y, respectively:

$\begin{matrix}{{{\psi_{1}\left( {x,y} \right)} = \frac{\partial{\theta\left( {x,y} \right)}}{\partial x}},{{\psi_{2}\left( {x,y} \right)} = \frac{\partial{\theta\left( {x,y} \right)}}{\partial y}},} & (2.3)\end{matrix}$where θ(x, y) is a 2D smoothing function that is well localized (aroundx=y=0) and isotropic. For any function ƒ(x, y) that exists in L²(R) thewavelet transform has two components with respect to ψ₁ and ψ₂ and canbe expressed in vectorial form as:

$\begin{matrix}{{{T_{\psi}\lbrack f\rbrack}\left( {b,a} \right)} = {\begin{pmatrix}{{T_{\psi\; 1}\lbrack f\rbrack} = {a^{2}{\int{d^{2}x\;{\psi_{1}\left( {a^{1}\left( {x - b} \right)} \right)}{f(x)}}}}} \\{{T_{\psi\; 2}\lbrack f\rbrack} = {a^{- 2}{\int{d^{2}x\;{\psi_{2}\left( {a^{- 1}\left( {x - b} \right)} \right)}{f(x)}}}}}\end{pmatrix}.}} & (2.4)\end{matrix}$

-   After performing an integration by parts, one can obtain    T _(ϕ)[ƒ](b, a)=a ⁻² ∇{∫d ² xϕ(a ⁻¹(x−b))ƒ(x)}  (2.5)    =∇{T _(ϕ)[ƒ](b, a)}  (2.6)    =∇{ϕ_(b,a)*ƒ}.   (2.7)-   We will take ϕ(x) to be the Gaussian function:

$\begin{matrix}{{\phi\left( {x,y} \right)} = e^{\frac{- {({x^{2} + y^{2}})}}{2},}} & (2.8)\end{matrix}$

If ∅(x ) is simply a smoothing function like the Gaussian, then equation2.7 amounts to define the 2D wavelet transform as the gradient vector off(x) smoothed by dilated versions of ∅(a⁻¹x) of this filter.

We will be referring to the wavelet-transform in terms of the modulusand argument:T _(ψ)[ƒ](b, a)=(

_(ψ)[ƒ](b, a),

_(ψ)[ƒ](b, a)).   (2.9)where

_(ψ)[ƒ](b, a)=[(T _(ψ) ₁ [ƒ](b, a))² +T _(ψ) ₂ [ƒ](b, a))²]^(1/2)  (2.10)and

_(ψ)[ƒ](b, a)=(T _(ψ) ₁ [ƒ](b, a)+iT _(ψ) ₂ [ƒ](b, a)).   (2.11)

As originally proposed by Mallat and collaborators, a very efficient wayto perform point-wise regularity analysis is to examine the wavelettransform modulus maxima. At any given scale a, the wavelet transformmodulus maxima (WTMM) are defined as the points b where the wavelettransform modulus, M [f](b, a), is locally maximum along the gradientdirection given by the wavelet transform argument, A [f](b, a). TheseWTMM, are inflection points of f *∅_(a)(x) and lie on connected chainscalled maxima chains. One only needs to record the position of the localmaxima of M along the maxima chains and the values of M [f] and A [f] atthe corresponding location. At each scale a, the wavelet analysisreduces to record the WTMM maxima (WTMMM) only. They indicate locallythe direction where the signal has the sharpest variation and aredisposed along connected curves across scales named maxima lines. Thewavelet transform skeleton will be defined as the set of maxima linesthat converge to the (x; y)-plane in the limit a→0+.

Discussion of Two Dimensional WTMM Method Mechanics

The 2D WTMM method relies upon the space-scale partitioning given by theWT skeleton. Let us define L(a) as the set of maxima lines that exist atthe scale a and which contain maxima at any scale a′≤a. The WTMM methodconsists in defining the following partition functions directly from theWTMMM that belong to the wavelet transform skeleton:

$\begin{matrix}{{{\left( {q,a} \right)} = {\left( {{M_{\psi}\lbrack f\rbrack}\left( {x,a^{\prime}} \right)} \right)^{q}}},} & (2.12)\end{matrix}$where q ∈ R. Compared to the classical box-counting techniques, theanalyzing wavelet, ψ, plays the role of a generalized “oscillating box”,the scale, a, defines its size, while the WT skeleton indicates how toposition our boxes to obtain a partition at the considered scale. Fromthe analogy that links the multifractal formalism to thermodynamics, onecan define the exponent τ(q) from the power law behavior of thepartition function:

(q, a)

a^(τ(q)), a→0⁺  (2.13)

Here, q and τ(q) are the inverse temperature and the free energyrespectively. This formalism replaces these variables with the Holderexponent, h, and the singularity spectrum, D(h). This means that theD(h) singularity spectrum of ƒ can be determined from the Legendretransform of the partition function scaling exponent τ(q):

$\begin{matrix}{{D(h)} = {\min\limits_{q}{\left( {{qh} - {\tau(q)}} \right).}}} & (2.14)\end{matrix}$

From the properties of the Legendre transform, the homogeneous fractalfunctions that involve singularities of unique Holder exponent

$h = \frac{\partial\tau}{\partial q}$are characterized by τ(q) spectrum which is a linear function of q. Anonlinear τ(q) curve is the signature of nonhomogeneous functions thatdisplay multifractal properties in the sense that the Holder exponenth(x) is a fluctuating quantity that depends upon the spatial position x.

The computation of the D(h) singularity spectrum, by the Legendretransform, requires first a smoothing of the τ(q) curve. This smoothingprocess may loss any interesting physics of phase transitions in thescaling properties of fractal functions. One can avoid directlyperforming the Legendre transform by considering the quantities h andD(h) as mean quantities defined with respect to their Boltzmann weightscomputed from the WTMMM:

W ψ ⁡ [ f ] ⁢ ( q , , a ) =  ⁢ ψ ⁡ [ f ] ⁢ ( x , a ′ )  q ⁢ ( q , a ) , (2.15 )where

(q, a) is the partition function. then one computes the expectationvalues:

h ⁡ ( q , a ) = ⁢ ln ⁢  ⁢ ψ ⁡ [ f ] ⁢ ( x , a ′ )  ⁢ ψ ⁡ [ f ] ⁢ ( q , , a ) ,( 2.16 )and

D ⁡ ( q , a ) = ⁢ ψ ⁡ [ f ] ⁢ ( q , , a ) ⁢ ln ⁡ ( ψ ⁡ [ f ] ⁢ ( q , , a ) ) (2.17 )from which one extracts

$\begin{matrix}{{h(q)} = {\lim\limits_{a->0^{+}}{{{h\left( {q,a} \right)}/\ln}\; a}}} & (2.18) \\{{D(q)} = {\lim\limits_{a->0^{+}}{{{D\left( {q,a} \right)}/\ln}\; a}}} & (2.19)\end{matrix}$and therefore the D(h) singularity spectrum.

It is important to note that while several provided methods use a twodimensional analysis, it is contemplated as within the scope of thepresent invention that three dimensional analyses may also beaccomplished. For example, provided methods may be used to analyze athree dimensional image generated via tomosynthesis, fluorescencemicroscopy, computed tomography, magnetic resonance imaging, or otherdigital methodology.

In some embodiments, the roughness analysis of tissue is not performedwith two dimensional images. The WTMM method as well as all othersimilar space-scale techniques that allow the calculation of the Hurstexponent (or any analogous exponent), such as, but not limited towavelet leaders, Fourier techniques, etc are also directly applicable to3D data cubes. Therefore, the assessment of cancerous or pre-canceroustissue regions through a roughness analysis is applicable to 3D data.For 3D data the distance between the estimated Hurst exponent (or otheranalogous exponent) and the critical value 0.5 is the same as for the 2Dcase. The strategy of analyzing square sub-regions neighboring a tumoror suspicious tissue area such as presented in FIG. 6 is directlygeneralizable to the analysis of cubic sub-regions in 3D.

Detecting Microcalcifications Through Wavelet Transform SkeletonSegmentation

In addition to determination of roughness exponents, the presentinvention provides methods and systems for detecting microcalcifications(MC) in subject tissue and characterizing their fractal dimension. Insome embodiments, provided methods use two dimensional (or, in someembodiments, three dimensional) WTMM to provide this information, thoughany mathematically related and/or similar multi-scale densityfluctuation assessment method yielding an exponent or spectrum ofexponents that is similar and/or can complement the use of the Hurstexponent and/or fractal dimension may be used.

The presence ofMC is one ofthe most important and sometimes the onlysign of cancer, for example, breast cancer in a screening mammogram. MCstypically appear as bright spots on mammograms and the number, size,type (Table 2) and distribution (Table 3) provide the radiologist withinformation about the potential malignancy of the lesion and allow anassessment score (Table 4) to be assigned to each case.

TABLE 2 Calcification Types Type Definition Pleomorphic Varying in shapeand size, smaller than 0.5 mm Punctate Round 0.5-1 mm Amorphous Small,round and hazy Fine Linear Branching Thin and linear or curved. Smallerthan 0.5 mm

TABLE 3 Calcification Distributions Distribution Definition LinearArranged in a line, suggesting being deposited in a duct SegmentalDeposits in ducts and branches of a segment or lobe Clustered At least 5calcifications in a small volume of tissue

TABLE 4 BI-RADS Assessment Score Score Interpretation 0 Cannot bedetermined by mammogram 1 No calcifications are present 2 No evidence ofmalignancy 3 Has less than 2% chance of being malignant 4 Consideredsuspicious 5 Has greater than or equal to 95% chance of being malignant6 Shown to be malignant through biopsy

By using the information listed in the above tables, radiologists canpredict the potential malignancy of the identified tumor on themammogram. For example, tumors that contain pleomorphic or fine linearbranching calcification types have a higher risk of being malignant thantumors that are classified as punctate. However, it is not uncommon fortumors to be made up of components that make it difficult forradiologists to the identify the tumor as cancerous or a benign diseasesuch as fibrocystic breast condition or a blunt injury. One benefit of aCAD is the objectivity of the software of having a range of indicesassociated with benign and/or malignant tumors.

Identifying Microcalcifications (MC)

In some embodiments, the present invention provides computer aideddiagnostic tools, using the WT methodology to detect MC by inspectingthe WT maxima chains. At the smallest scale resolved by the WTmicroscope, 7 pixels, MC which can be considered at strong singularitiesare contour-shaped by some maxima chains. Since the average size of MCis about 200 μm, or 5 pixels, these singularities are identified by themathematical microscope as Dirac singularities. Thus, the correspondingmaxima lines pointing to the MC are likely to display scaling propertieswith a Holder exponent h=−1 down to scales of the order of the MC sizewhere one can observe a cross-over to the value h=0 as the signature ofthe MC boundary. Therefore, one can perform a classification of theselines according to the behavior of the M_(ψ) [f] and then separate MC,h=−1, from dense tissue, h˜0.65, and fatty tissue, h˜0.30.

As pointed out above, the MC wavelet-transform skeleton can be used tocompute the partition functions, and thus fully characterize the fractalgeometry of the MC cluster.

Separating the MC from the subject tissue (e.g., breast tissue) in thisway is key, since it allows one to consider the properties of thebackground tissue separate from the properties of the MC. Using providedmethods, one is able to quantify the arrangement and organization of theMC and consider the distribution of the tumor, making the similarinformation currently used in Table 3 less subjective. At the same time,one can classify the tissue and analyze any interesting physical orstructural changes from previous mammograms or surrounding tissue.Therefore, the two dimensional WTMM method has the ability to be apowerful CAD. The information obtained from the wavelet-transformanalysis can help quantify data that would otherwise be consideredsubjective. In some embodiments, provided methods use this informationto aid in the development of an objective mammographic classification,which can be used to judge the potential malignancy of aradiologist-identified suspicious region.

Assessment of Three Dimensional Fractal Geometry from Multiple TwoDimensional Views

In some embodiments, characterization of the fractal geometry of MCclusters from a plurality of separate two-dimensional views of the sameregion of tissue (e.g. a breast), allows for an assessment of theoverall fractal dimension of the tissue region of interest, such asthrough application of the WT methods described herein. In other words,according to various embodiments, by using a plurality of separatetwo-dimensional views of the same region of tissue one can assess thefractal dimension of the three dimensional tissue region of interest.These concepts are described in more detail in the Examples below.

In some embodiments, non-cancerous or non-pre-cancerous tissue will havea Euclidean fractal dimension (i.e. an integer fractal dimension). Insome embodiments, a cancerous or pre-cancerous tissue will have anon-integer fractal dimension indicating an invasive morphology.

In some embodiments, the present invention also provides methodsincluding providing a first view of a region of tissue, providing asecond view of a region of tissue, calculating a first fractal dimensionfor the first view of the region of tissue, and calculating a secondfractal dimension for the second view of the region of tissue, whereinif the fractal dimension of at least one of the first fractal dimensionand the second fractal dimension is in the fractal zone, the region oftissue is considered cancerous. In some embodiments, provided methodsfurther comprise treating the region of tissue if it is cancerous orpre-cancerous.

In some embodiments, the “fractal zone” is defined as a polygonconsisting of a central square and a first, second, third, and fourthextending triangular region as plotted on a graph of the fractaldimension of the first view of the region of tissue by the fractaldimension of the second view of the region of tissue.

In order to provide greater clarity around the concept of fractal zones,several non-limiting exemplary embodiments are provided. In someembodiments, such as that shown in FIG. 11, a fractal zone may bedefined as consisting of a central square and a first, second, third,and fourth extending triangular region as plotted on a graph of thefractal dimension of the first view of the region of tissue by thefractal dimension of the second view of the region of tissue with thefollowing dimensions:

central square: (1.2, 1.2), (1.2, 1.8), (1.8, 1.2)(1.8, 1.8);

first extending triangular region: (0.5, 1.5), (1.2, 1.2), (1.2, 1.8);

second extending triangular region: (1.5, 0.5), (1.2, 1.2), (1.8, 1.2);

third extending triangular region: (1.5, 2.3), (1.2, 1.8), (1.8, 1.8);and

fourth extending triangular region: (2.3, 1.5), (1.8, 1.2), (1.8, 1.8).

In some embodiments, a fractal zone may be defined as consisting of acentral square and a first, second, third, and fourth extendingtriangular region as plotted on a graph of the fractal dimension of thefirst view of the region of tissue by the fractal dimension of thesecond view of the region of tissue with the following dimensions:

central square: (1.1, 1.1), (1.1, 1.9), (1.9, 1.1)(1.9, 1.9);

first extending triangular region: (0.5, 1.5), (1.1, 1.1), (1.1, 1.9);

second extending triangular region: (1.5, 0.5), (1.1, 1.1), (1.9, 1.1);

third extending triangular region: (1.5, 2.3), (1.1, 1.9), (1.9, 1.9);and

fourth extending triangular region: (2.3, 1.5), (1.9, 1.1), (1.9, 1.9).

In some embodiments, a fractal zone may be defined as consisting of acentral square and a first, second, third, and fourth extendingtriangular region as plotted on a graph of the fractal dimension of thefirst view of the region of tissue by the fractal dimension of thesecond view of the region of tissue with the following dimensions:

central square: (1.3, 1.3), (1.3, 1.7), (1.7, 1.3)(1.7, 1.7);

first extending triangular region: (0.5, 1.5), (1.3, 1.3), (1.3, 1.7);

second extending triangular region: (1.5, 0.5), (1.3, 1.3), (1.7, 1.3);

third extending triangular region: (1.5, 2.3), (1.3, 1.7), (1.7, 1.7);and

fourth extending triangular region: (2.3, 1.5), (1.7, 1.3), (1.7, 1.7).

It is contemplated that the precise boundaries of the fractal zone mayvary according to tissue type and/or tumor type (or subtype).Additionally, without wishing to be held to a particular theory it ispossible that the precise borders of the fractal zone may change overtime as tissue changes from normal tissue to pre-cancerous or canceroustissue. One of skill in the art will be able to ascertain from thepresent disclosure how to determine the appropriate boundaries of thefractal zone in a particular clinical scenario using no more thanroutine experimentation.

In some embodiments, provided methods may be applied to more than twoviews of a tissue region of interest. In some embodiments, providedmethods may be applied to three, four, five, six, seven, eight, nine,ten, fifteen, twenty or more views of a particular tissue region ofinterest. One of skill will understand how the addition of additionaltwo-dimension images to the analysis of provided methods will alter thedefinition of the fractal zone. For example, if three views of a tissueregion of interest are used, the fractal zone may then be defined as theunion of seven juxtaposed regions: one central cube and six extendingpyramidal or conical regions. In some embodiments, wherein four or moreviews of a tissue region of interest are used, it may be more convenientto define the fractal zone purely mathematically, rather than in termsof geometric shapes. Additionally, the borders of the fractal zone,though described herein (including in the below Examples) as lines, maycomprise other forms and definitions. For example, in some embodiments,the borders of the fractal zone may be smooth curves, jagged lines, oreven fractals.

In addition to the use of the WTMM method to calculate the fractaldimension of clusters of microcalcifications , other methods may also beused according to various embodiments. In some embodiments, any fractaltechnique or techniques yielding a fractal dimension, such as, but notlimited to: box-counting techniques, the perimeter-area relationship,packing dimension, Hausdorff dimension, capacity dimension, correlationdimension, the generalized dimensions (multifractal), and/or anyspace-scale technique that yields a value quantifying the structuralcomplexity of a tumor or object of interest as a function of scale maybe used.

As described elsewhere herein, provided methods may be applied to anytissue that may contain or comprise cancer. Non-limiting exemplary suchtissues include breast tissue, brain tissue, lung tissue, kidney tissue,liver tissue, uterine tissue, dermal tissue, and pancreatic tissue.

Treatment of Cancerous or Pre-Cancerous Tissue

According to various embodiments, provided methods may be used to detectand/or characterize the presence of cancerous or pre-cancerous tissue ina subject. It is contemplated that if cancerous or pre-cancerous tissueis discovered, that treatment of such tissue would commence inaccordance with sound medical judgment. It is further contemplated thattreatment may occur with any anti-cancer therapy, whether currentlyknown or discovered in the future.

While any anti-cancer therapy may be appropriate for use in someembodiments, exemplary types of anti-cancer therapy are described belowin order to better illustrate some of the principles of the presentinvention.

Traditional therapies or anticancer agents include surgery, radiotherapy(γ-radiation, neutron beam radiotherapy, electron beam radiotherapy,proton therapy, brachytherapy, and systemic radioactive isotopes, toname a few), endocrine therapy, biologic response modifiers(interferons, interleukins, and tumor necrosis factor (TNF) to name afew), hyperthermia and cryotherapy, agents to attenuate any adverseeffects (e.g., antiemetics), and other approved chemotherapeutic drugs,including, but not limited to, alkylating drugs (mechlorethamine,chlorambucil, Cyclophosphamide, Melphalan, Ifosfamide), antimetabolites(Mcthotrcxatc), purinc antagonists and pyrimidinc antagonists(6-Mercaptopurine, 5-Fluorouracil, Cytarabile, Gemcitabine), spindlepoisons (Vinblastine, Vincristine, Vinorelbine, Paclitaxel),podophyllotoxins (Etoposide, Irinotecan, Topotecan), antibiotics(Doxorubicin, Bleomycin, Mitomycin), nitrosoureas (Carmustine,Lomustine), inorganic ions (Cisplatin, Carboplatin), enzymes(Asparaginase), and hormones (Tamoxifen, Leuprolide, Flutamide, andMegestrol), to name a few. Any and all of these exemplary therapies, aswell as other cancer therapies known by one of skill in the art, may beused in connection with the present invention.

It is contemplated that the specific treatment, including dose, dosingregimen, mode of administration, and timing of the onset and terminationof therapy will be determined by a medical practitioner in accordancewith sound medical judgment.

Systems and Devices for Implementing Provided Methods

The present invention also provides systems and devices, such ascomputing devices, implementing provided methods. In some embodiments, acomputing device maybe a cloud computing device, a mobile computingdevice, or any other application-appropriate computing device.

As shown in FIG. 13, an exemplary implementation of a networkenvironment 1300 for use with provided methods is shown and described.In brief overview, referring now to FIG. 13, a block diagram of anexemplary cloud computing environment 1300 is shown and described. Thecloud computing environment 1300 may include one or more resourceproviders 1302 a, 1302 b, 1302 c (collectively, 1302). Each resourceprovider 1302 may include computing resources. In some implementations,computing resources may include any hardware and/or software used toprocess data. For example, computing resources may include hardwareand/or software capable of executing algorithms, computer programs,and/or computer applications. In some implementations, exemplarycomputing resources may include application servers and/or databaseswith storage and retrieval capabilities. Each resource provider 1302 maybe connected to any other resource provider 1302 in the cloud computingenvironment 1300. In some implementations, the resource providers 1302may be connected over a computer network 1308. Each resource provider1302 may be connected to one or more computing device 1304 a, 1304 b,1304 c (collectively, 1304), over the computer network 1308.

The cloud computing environment 1300 may include a resource manager1306. The resource manager 1306 may be connected to the resourceproviders 1302 and the computing devices 1304 over the computer network1308. In some implementations, the resource manager 1306 may facilitatethe provision of computing resources by one or more resource providers1302 to one or more computing devices 1304. The resource manager 1306may receive a request for a computing resource from a particularcomputing device 1304. The resource manager 1306 may identify one ormore resource providers 1302 capable of providing the computing resourcerequested by the computing device 1304. The resource manager 1306 mayselect a resource provider 1302 to provide the computing resource. Theresource manager 1306 may facilitate a connection between the resourceprovider 1302 and a particular computing device 1304. In someimplementations, the resource manager 1306 may establish a connectionbetween a particular resource provider 1302 and a particular computingdevice 1304. In some implementations, the resource manager 1306 mayredirect a particular computing device 1304 to a particular resourceprovider 1302 with the requested computing resource.

FIG. 14 shows an example of a computing device 1400 and a mobilecomputing device 1450 that can be used to implement the techniquesdescribed in this disclosure. The computing device 1400 is intended torepresent various forms of digital computers, such as laptops, desktops,workstations, personal digital assistants, servers, blade servers,mainframes, and other appropriate computers. The mobile computing device1450 is intended to represent various forms of mobile devices, such aspersonal digital assistants, cellular telephones, smart-phones, andother similar computing devices. The components shown here, theirconnections and relationships, and their functions, are meant to beexamples only, and are not meant to be limiting.

The computing device 1400 includes a processor 1402, a memory 1404, astorage device 1406, a high-speed interface 1408 connecting to thememory 1404 and multiple high-speed expansion ports 1410, and alow-speed interface 1412 connecting to a low-speed expansion port 1414and the storage device 1406. Each of the processor 1402, the memory1404, the storage device 1406, the high-speed interface 1408, thehigh-speed expansion ports 1410, and the low-speed interface 1412, areinterconnected using various busses, and may be mounted on a commonmotherboard or in other manners as appropriate. The processor 1402 canprocess instructions for execution within the computing device 1400,including instructions stored in the memory 1404 or on the storagedevice 1406 to display graphical information for a GUI on an externalinput/output device, such as a display 1416 coupled to the high-speedinterface 1408. In other implementations, multiple processors and/ormultiple buses may be used, as appropriate, along with multiple memoriesand types of memory. Also, multiple computing devices may be connected,with each device providing portions of the necessary operations (e.g.,as a server bank, a group of blade servers, or a multi-processorsystem).

The memory 1404 stores information within the computing device 1400. Insome implementations, the memory 1404 is a volatile memory unit orunits. In some implementations, the memory 1404 is a non-volatile memoryunit or units. The memory 1404 may also be another form ofcomputer-readable medium, such as a magnetic or optical disk.

The storage device 1406 is capable of providing mass storage for thecomputing device 1400. In some implementations, the storage device 1406may be or contain a computer-readable medium, such as a floppy diskdevice, a hard disk device, an optical disk device, or a tape device, aflash memory or other similar solid state memory device, or an array ofdevices, including devices in a storage area network or otherconfigurations. Instructions can be stored in an information carrier.The instructions, when executed by one or more processing devices (forexample, processor 1402), perform one or more methods, such as thosedescribed above. The instructions can also be stored by one or morestorage devices such as computer- or machine-readable mediums (forexample, the memory 1404, the storage device 1406, or memory on theprocessor 1402).

The high-speed interface 1408 manages bandwidth-intensive operations forthe computing device 1400, while the low-speed interface 1412 manageslower bandwidth-intensive operations. Such allocation of functions is anexample only. In some implementations, the high-speed interface 1408 iscoupled to the memory 1404, the display 1416 (e.g., through a graphicsprocessor or accelerator), and to the high-speed expansion ports 1410,which may accept various expansion cards (not shown). In theimplementation, the low-speed interface 1412 is coupled to the storagedevice 1406 and the low-speed expansion port 1414. The low-speedexpansion port 1414, which may include various communication ports(e.g., USB, Bluetooth®, Ethernet, wireless Ethernet) may be coupled toone or more input/output devices, such as a keyboard, a pointing device,a scanner, or a networking device such as a switch or router, e.g.,through a network adapter.

The computing device 1400 may be implemented in a number of differentforms, as shown in the figure. For example, it may be implemented as astandard server 1420, or multiple times in a group of such servers. Inaddition, it may be implemented in a personal computer such as a laptopcomputer 1422. It may also be implemented as part of a rack serversystem 1424. Alternatively, components from the computing device 1400may be combined with other components in a mobile device (not shown),such as a mobile computing device 1450. Each of such devices may containone or more of the computing device 1400 and the mobile computing device1450, and an entire system may be made up of multiple computing devicescommunicating with each other.

The mobile computing device 1450 includes a processor 1452, a memory1464, an input/output device such as a display 1454, a communicationinterface 1466, and a transceiver 1468, among other components. Themobile computing device 1450 may also be provided with a storage device,such as a micro-drive or other device, to provide additional storage.Each of the processor 1452, the memory 1464, the display 1454, thecommunication interface 1466, and the transceiver 1468, areinterconnected using various buses, and several of the components may bemounted on a common motherboard or in other manners as appropriate.

The processor 1452 can execute instructions within the mobile computingdevice 1450, including instructions stored in the memory 1464. Theprocessor 1452 may be implemented as a chipset of chips that includeseparate and multiple analog and digital processors. The processor 1452may provide, for example, for coordination of the other components ofthe mobile computing device 1450, such as control of user interfaces,applications run by the mobile computing device 1450, and wirelesscommunication by the mobile computing device 1450.

The processor 1452 may communicate with a user through a controlinterface 1458 and a display interface 1456 coupled to the display 1454.The display 1454 may be, for example, a TFT (Thin-Film-Transistor LiquidCrystal Display) display or an OLED (Organic Light Emitting Diode)display, or other appropriate display technology. The display interface1456 may comprise appropriate circuitry for driving the display 1454 topresent graphical and other information to a user. The control interface1458 may receive commands from a user and convert them for submission tothe processor 1452. In addition, an external interface 1462 may providecommunication with the processor 1452, so as to enable near areacommunication of the mobile computing device 1450 with other devices.The external interface 1462 may provide, for example, for wiredcommunication in some implementations, or for wireless communication inother implementations, and multiple interfaces may also be used.

The memory 1464 stores information within the mobile computing device1450. The memory 1464 can be implemented as one or more of acomputer-readable medium or media, a volatile memory unit or units, or anon-volatile memory unit or units. An expansion memory 1474 may also beprovided and connected to the mobile computing device 1450 through anexpansion interface 1472, which may include, for example, a SIMM (SingleIn Line Memory Module) card interface. The expansion memory 1474 mayprovide extra storage space for the mobile computing device 1450, or mayalso store applications or other information for the mobile computingdevice 1450. Specifically, the expansion memory 1474 may includeinstructions to carry out or supplement the processes described above,and may include secure information also. Thus, for example, theexpansion memory 1474 may be provided as a security module for themobile computing device 1450, and may be programmed with instructionsthat permit secure use of the mobile computing device 1450. In addition,secure applications may be provided via the SIMM cards, along withadditional information, such as placing identifying information on theSIMM card in a non-hackable manner.

The memory may include, for example, flash memory and/or NVRAM memory(non-volatile random access memory), as discussed below. In someimplementations, instructions are stored in an information carrier and,when executed by one or more processing devices (for example, processor1452), perform one or more methods, such as those described above. Theinstructions can also be stored by one or more storage devices, such asone or more computer- or machine-readable mediums (for example, thememory 1464, the expansion memory 1474, or memory on the processor1452). In some implementations, the instructions can be received in apropagated signal, for example, over the transceiver 1468 or theexternal interface 1462.

The mobile computing device 1450 may communicate wirelessly through thecommunication interface 1466, which may include digital signalprocessing circuitry where necessary. The communication interface 1466may provide for communications under various modes or protocols, such asGSM voice calls (Global System for Mobile communications), SMS (ShortMessage Service), EMS (Enhanced Messaging Service), or MMS messaging(Multimedia Messaging Service), CDMA (code division multiple access),TDMA (time division multiple access), PDC (Personal Digital Cellular),WCDMA (Wideband Code Division Multiple Access), CDMA2000, or GPRS(General Packet Radio Service), among others. Such communication mayoccur, for example, through the transceiver 1468 using aradio-frequency. In addition, short-range communication may occur, suchas using a Bluetooth®, Wi-Fi™, or other such transceiver (not shown). Inaddition, a GPS (Global Positioning System) receiver module 1470 mayprovide additional navigation- and location-related wireless data to themobile computing device 1450, which may be used as appropriate byapplications running on the mobile computing device 1450.

The mobile computing device 1450 may also communicate audibly using anaudio codec 1460, which may receive spoken information from a user andconvert it to usable digital information. The audio codec 1460 maylikewise generate audible sound for a user, such as through a speaker,e.g., in a handset of the mobile computing device 1450. Such sound mayinclude sound from voice telephone calls, may include recorded sound(e.g., voice messages, music files, etc.) and may also include soundgenerated by applications operating on the mobile computing device 1450.

The mobile computing device 1450 may be implemented in a number ofdifferent forms, as shown in the figure. For example, it may beimplemented as a cellular telephone 1480. It may also be implemented aspart of a smart-phone 1482, personal digital assistant, or other similarmobile device.

Various implementations of the systems and techniques described here canbe realized in digital electronic circuitry, integrated circuitry,specially designed ASICs (application specific integrated circuits),computer hardware, firmware, software, and/or combinations thereof Thesevarious implementations can include implementation in one or morecomputer programs that are executable and/or interpretable on aprogrammable system including at least one programmable processor, whichmay be special or general purpose, coupled to receive data andinstructions from, and to transmit data and instructions to, a storagesystem, at least one input device, and at least one output device.

These computer programs (also known as programs, software, softwareapplications or code) include machine instructions for a programmableprocessor, and can be implemented in a high-level procedural and/orobject-oriented programming language, and/or in assembly/machinelanguage. As used herein, the terms machine-readable medium andcomputer-readable medium refer to any computer program product,apparatus and/or device (e.g., magnetic discs, optical disks, memory,Programmable Logic Devices (PLDs)) used to provide machine instructionsand/or data to a programmable processor, including a machine-readablemedium that receives machine instructions as a machine-readable signal.The term machine-readable signal refers to any signal used to providemachine instructions and/or data to a programmable processor.

To provide for interaction with a user, the systems and techniquesdescribed here can be implemented on a computer having a display device(e.g., a CRT (cathode ray tube) or LCD (liquid crystal display) monitor)for displaying information to the user and a keyboard and a pointingdevice (e.g., a mouse or a trackball) by which the user can provideinput to the computer. Other kinds of devices can be used to provide forinteraction with a user as well; for example, feedback provided to theuser can be any form of sensory feedback (e.g., visual feedback,auditory feedback, or tactile feedback); and input from the user can bereceived in any form, including acoustic, speech, or tactile input.

The systems and techniques described here can be implemented in acomputing system that includes a back end component (e.g., as a dataserver), or that includes a middleware component (e.g., an applicationserver), or that includes a front end component (e.g., a client computerhaving a graphical user interface or a Web browser through which a usercan interact with an implementation of the systems and techniquesdescribed here), or any combination of such back end, middleware, orfront end components. The components of the system can be interconnectedby any form or medium of digital data communication (e.g., acommunication network). Examples of communication networks include alocal area network (LAN), a wide area network (WAN), and the Internet.

The computing system can include clients and servers. A client andserver are generally remote from each other and typically interactthrough a communication network. The relationship of client and serverarises by virtue of computer programs running on the respectivecomputers and having a client-server relationship to each other.

In view of the structure, functions and apparatus of the systems andmethods described here, in some implementations, systems and methods forcharacterizing potentially cancerous and/or pre-cancerous tissue areprovided. Having described certain implementations of methods andapparatus for provided methods, it will now become apparent to one ofskill in the art that other implementations incorporating the conceptsof the disclosure may be used. Therefore, the disclosure should not belimited to certain implementations, but rather should be limited only bythe spirit and scope of the following claims.

Throughout the description, where apparatus and systems are described ashaving, including, or comprising specific components, or where processesand methods are described as having, including, or comprising specificsteps, it is contemplated that, additionally, there are apparatus, andsystems of the disclosed technology that consist essentially of, orconsist of, the recited components, and that there are processes andmethods according to the disclosed technology that consist essentiallyof, or consist of, the recited processing steps.

It should be understood that the order of steps or order for performingcertain action is immaterial so long as the disclosed technology remainsoperable. Moreover, two or more steps or actions may be conductedsimultaneously.

EXAMPLES Example 1 Characterization of Breast Tumor Organization

This Example shows that the 2D WTMM method has the ability tocharacterize breast tumors and their microenvironment. Data was obtainedfrom the Digital Database for Screening Mammography (DDSM), describedabove. Cases having a suspicious region, i.e. containing a benign ormalignant tumor, were considered. From these cases, only cases havingone set of microcalcifications were kept for further investigation andmasses were disregarded for this analysis. Using the methods describedabove, we were able to fully characterize the lesion by the fractaldimension of the breast tumor and the roughness of the microenvironmentgiven by the Hurst exponent.

In this Example, a total of 128 images, 78 of which are benign and 50 ofwhich are malignant, were analyzed. Since each mammographic casecontains two images, one corresponding to the mediolateral oblique (MLO)view, and the other to the cranio-caudal (CC) view with theirprojections shown in FIG. 1, we combined the information to reconstructan estimate of the 3D structure of the tumor embedded into the breasttissue. The information was combined by creating a scatter plot of theminimum fractal dimension versus the maximum fractal dimension as shownin FIG. 2.

The Mclust function in R was applied to the two data sets, the benignand malignant scatter plots. The results from the function shown in FIG.3, suggests that benign tumors are constructed from three subpopulationsand malignant tumors are constructed from two subpopulations. Here, itis important to note the centers of the components identified by thefunction. Benign breast lesions' three components are centered at(0.84,1.04), (0.85,1.78) and (1.78,1.88), while malignant breastlesions' two components are centered at (1.07,1.62) and (1.49,1.52).

The center of the benign categories demonstrate how these tumors arenon-invasive since the populations fractal dimensions lay in Euclideanspace and are represented by lines, sheets and spheres. However, bothmalignant categories are shown by invasive fractal dimensions andrepresented by fractal structures in breast tissue.

Example 2 Classification of Tumor Prone Tissue

This example illustrates how provided methods may be used to betterunderstand the mechanisms that drive the differing organization ofbenign and malignant tumors, and specifically the microenvironment of aradiologist-identified suspicious region.

To gain a deeper understanding of the behavior and development of breastlesions, neighboring tissue was classified according to the twodimensional WTMM methods described above to survey the microenvironmentof the tumors (also see Kestener et al., Wavelet-based multifractalformalism to assist in diagnosis in digitized mammograms, 2001, ImageAnal Stereol, 20: 169-174 for an application of this methodology).Briefly, the 2D WTMM method was used to analyze mammary parenchyma. Thisanalysis was used to generate a Hurst exponent (H) as a measure of theroughness of a sample. The Hurst score was used to classify breasttissue as fatty (H=0.25±0.05), dense (H=0.65±0.05).

To categorize the neighboring tissue, each image of a breast lesion wassegmented into 9 subimages, corresponding to the same size and shape ofthe radiologist encircled tumor. Once the images were properlysegmented, only the central part of the neighboring images were analyzedto counter any edge effects that may disrupt the analysis. The roughsurface images were analyzed via the above methodology.

As shown in FIG. 4, the results allowed for the characterization ofabnormal tissue corresponding to a benign tumor and malignant tumor. InFIG. 4, blue represents H=(0-0.45), or fatty tissue, red representsH=(0.55-1), or dense tissue, and yellow represents H=(0.45; 0.55). It isimportant to note that in the cancer sample shown in FIG. 4 there areseveral yellow regions identified to have H=(0.45-0.55), where in benignand healthy samples, no such tissue classification was found to exist.

In order to statistically determine if the microenvironments of benignand malignant breast lesions are significantly different, we constructeda t-test to obtain a p-value. Our hypothesis is that themicroenvironment of malignant breast tumors are categorized as H=0.5,while benign tumors have a microenvironment categorized as H=0.3 orH=0.65, corresponding to healthy fatty or dense tissue, respectively. Weobtained one value per case by averaging the calculating |H −0.5| valuesof the neighboring tissue for both the CC and MLO views. From here, theH values were averaged for each case. By computing ΔH=<|H −0.5|>, we canexpect benign tissue to have a higher value characterizing themicroenvironment compared to malignant tissue.

The boxplot in FIG. 5 shows the distribution of ΔH for both benign andmalignant microenvironments. Here, the max and min values are shown onthe plot as 0.33 and 0.07 for benign and 0.26 and 0.08 for cancerrespectively. As shown in the boxplot, three outliers were identifiedfor benign and one for cancer, meaning these values for ΔH are 1.5 timesmore than the maximum ΔH value.

Since the boxplot suggests there may be some non-normality of the data,a test for normality was performed in order to decide if atransformation of the data would be necessary before proceeding to thet-test. The Shapiro-Wilk test, first published by S. Shapiro and M. Wilkin 1965, tests the null hypothesis that a sample comes from a normallydistributed population. The test statistic is given by:

$\begin{matrix}{{W = \frac{\left( {\sum\limits_{i = 1}^{n}{a_{i}x_{i}}} \right)^{2}}{\sum\limits_{i = 1}^{n}\left( {x_{i} - \overset{\_}{x}} \right)^{2}}},} & (5.1)\end{matrix}$where x is the sample mean, x_((i)) is the i order statistic, and

${a_{i} = \frac{m^{T}V^{- 1}}{\left( {m^{T}V^{- 1}V^{- 1}m} \right)^{(\frac{1}{2})}}},$where m=(m₁; m₂; . . . ; m_(n))_(T) and m_(i) is expected value of theorder statistic and V is the covariance matrix.

The result from performing the Shapiro-Wilk Test on our data shows thatthe data was not normally distributed since p=0.001 for benign andp=0.033 for cancer. A log transformation was conducted and the resultsin Table 5 demonstrates normal data.

TABLE 5 Results From Shapiro-Wilk Test Group W P-Value Log (BenignTissue) 0.97 0.49 Log (Cancer Tissue) 0.96 0.41

Once the data has been transfol led in order for the data sets to followa normal distribution, we checked to make sure the two data sets haveequal variances by using the F-ratio test. Here, the null hypothesis isthat two normal populations have the same variance. The test statisticis:

$\begin{matrix}{{F = \frac{S_{X}^{2}}{S_{Y}^{2}}},} & (5.2)\end{matrix}$where X and Y are independent and normal, and S² _(X) and S² _(Y) arethe sample variances. Thus, we would reject H₀ if F is too large or toosmall. The results from the F-ratio test in Table 6 tells us to rejectH₀.

TABLE 6 Results from F-Ratio Test F Numerator df Denominator df p-value1.04 38 24 0.93

Once we have checked the assumptions of the parametric test, we canperform the necessary t-test. We can define μ_(B) as ΔH for the benignmicroenvironment and μC as ΔH for the cancer microenvironment. We willbe testing the null hypothesis that μC≥μ_(B), making the alternativehypothesis μC<μB. The result, shown in Table 7 , was a p-value of 0.04.Thus, we can reject H₀ and conclude there is a significant difference inthe log(ΔH) values characterizing the microenvironments of benign andmalignant breast lesions.

TABLE 7 T-Test Results of Cancer and Benign ΔH t df p-value −1.79 52.110.04

It is key to point out that H=0.5 is not found on healthy mammograms inprevious studies and less often on the benign sample studied here.Another key piece is that H=0.5 corresponds to no correlation and a lossof structure. This suggests that the tissue composing the environment ofmalignant tumors has lost its structure and may even precede the onsetor progression of the microcalcifications (MC). Thought to be anindicator of the early development of breast cancer. Thus, this Exampleshows that calculation of a Hurst exponent according to provided methodsprovides a powerful tool in the early detection and diagnosis of breastcancer.

Example 3 Alternative Analysis of Prone Tissue

As indicated above in Example 2, obtaining one average ΔH value persubject by averaging the calculated |H −0.5| values of neighboringtissue for both the MLO and CC views provided valuable insight that theloss of microenvironmental structure is indicative of the development oftumors, with a higher level of disorder being found in more serious(malignant) tumors.

In this example, rather than calculating a single average ΔH value persubject (consisting of up to 16 neighbors per image, 8 in each of theMLO and CC views), the distribution of ΔH=|H −0.5| for each of theneighboring regions was taken individually. This methodology provides asignificantly larger sample size than the analysis in Example 2 becausethe sample size becomes the number of subjects x the number ofneighboring images around each lesion, for a total of (up to) 16ΔH=|H−0.5| values per subject. When the data in Example 2 is analyzedusing this method, the p-value between malignant and benign tissues isan even more significant p=0.001.

The analysis in this Example is likely to be of significant utility invarious embodiments because it is better able to account for variationswithin a subject and may be able to better define cancerous orpre-cancerous regions within a target tissue.

Example 4 Characterization of Tissue Microenvironment

In this Example, data from analyzed breast tissue was characterized on aper neighboring region basis, as opposed to a per patient basis. Forthis Example, a total of 1,131 images of tissue classified as malignant,1,131 images of tissue classified as benign, and 64 images of tissueclassified as normal were analyzed. Specifically, the number of regionsin analyzed microenvironments characterized as fatty, dense or disruptedwere calculated from the samples examined in Example 1. Table 8 showsthe results of this analysis:

TABLE 8 Classification of Tissue Microenvironment % of images % ofimages for Total for which which H > 0.45 % of images for number TissueH <= 0.45 and H < 0.55 which H >= 0.55 of Type (FATTY) (DISRUPTED)(DENSE) images Malignant 39.8% 22.6% 37.6% 100% Benign 47.9% 17.9% 34.2%100% Normal 51.6% 10.9% 37.5% 100% (Healthy)

As shown in Table 8, the percentage of disrupted regions was highest inmalignant tissue, followed by benign tissue, with normal tissue havingthe lowest levels of disruption across the tissue. Exemplary photographsof the differences in observed disruption may be found in FIG. 6. It isimportant to note that the percentage of disrupted regions for normalbreast tissue is not 0%. Without wishing to be held to a particulartheory, it is possible these disrupted regions may be representative ofa transitional state of the tissue that may be prone to the eventualdevelopment of a tumor. Longitudinal studies will help to determine thenature of disrupted regions in breast tissue classified as normal bycurrent diagnostic methodologies.

In order to more accurately calculate the proportion of disruptedregions in malignant, benign, and normal tissue (particularly given thelow number of normal tissue images analyzed above), a second study ofthe same design as that described above was performed. In the secondstudy a total of 1,081 malignant images, 1,122 benign images, and 740images classified as normal (more than ten times that number from thefirst study) were included. The results are shown below in Table 9:

TABLE 9 Classification of Tissue Microenvironment Normal Tissue BenignTissue Malignant Tissue Proportion 44/740 = 5.9% 202/1,122 = 18.0%256/1,081 = 23.7% (r) 95% (4.2%-7.6%)* (15.8%-20.3%)** (21.1%-26.2%)Confidence Interval *p = 9.4 × 10⁻¹⁴ as compared to benign tissue; p =2.2 × 10⁻¹⁶ as compared to malignant tissue **p = 0.0012 as compared tomalignant tissue

As shown in Table 9, using a larger sample size of tissue classified asnormal, the proportion of tissue exhibiting disrupted surroundingregions drops from 10.9% in Table 8 to 5.9% in Table 9. The lowerobserved proportion of images classified as normal exhibiting disruptedtissue further supports the use of roughness and disorder in surroundingtissue as an indicator of cancer or pre-cancer in a tissue, and possiblymalignancy. Without wishing to be held to a particular theory, it ispossible that tissue otherwise classified as normal that exhibits somedegree of disrupted tissue in the surrounding microenvironment may bepre-cancerous or have an increased likelihood of developing cancer inthe future as compared to tissue that does not exhibit disruption in thesurrounding microenvironment.

Example 5 Detection of Microcalcification (MC) Clusters and Calculationof their Fractal Dimension

Methods—Samples

In this Example, images that were analyzed were obtained from theDigital Database for Screening Mammography (DDSM) at the University ofSouth Florida. The databank contains over 2,600 studies made up ofnormal, benign, benign without call back and malignant mammograms allcategorized by an expert radiologist. Each study has two images of eachbreast, consisting of a mediolateral-oblique (MLO) view andcranio-caudal (CC) view with any suspicious region circled by aradiologist. The suspicious region could contain a mass and/ormicrocalcifications (MC), but only the cases that were classified ashaving exactly one tumor composed of only MC in the benign and malignantcategories were looked at in this particular study.

In addition to only considering tumors consisting of MC, anymammographic images that contained artifacts inside the radiologist'sencircled region were discarded due to the impact it has on theanalysis. These artifacts could include scratches, hair, deodorant,patient movement, scanner artifacts (rollers slipped), pacemaker, breastimplants, skin markers (for scars, moles, and nipples, as well as markedlumps of breast pain), metallic foreign bodies, and fingerprints. Some(but not all) of these effects were recorded under notes in the DDSMwebsite. In this Example, a total of 59 cases were considered,corresponding to 118 images of size greater than 2562 pixels, 34 ofwhich are benign (68 images) and 25 of which are malignant (50 images).

Methods—The 2D WTMM Method

In this Example, the two dimensional (2D) WTMM method is used tocharacterize images of breast tissue, as described above. Most of theexisting CAD methods, whether specifically designed for two dimensional(2D) mammograms or more recently, for three dimensional (3D) breasttomosynthesis, have been elaborated on the prerequisite that thebackground roughness fluctuations of normal breast texture arestatistically homogeneous and uncorrelated, which precludes theirability to adequately characterize background tissue. The majority ofthe fractal methods used to examine and classify mammographic breastlesions rely on the estimate of the Hurst exponent (or its variousmathematical equivalents), which globally characterizes the self-similarproperties of the landscape in question. However, the 2D WTMM methodtakes in account that the function defining the image may bemultifractal, therefore requiring the use of the Holder exponent(M=Ka^(h)) to characterize the local regularity at a particular point.

The 2D WTMM method requires us to define a smoothing function, ∅(x, y),in two dimensions that is a well-localized isotropic function around theorigin. In this Example, we used the Gaussian function and define thewavelets as:

${\psi_{1}\left( {x,y} \right)} = {{\frac{\partial{\phi\left( {x,y} \right)}}{\partial x}\mspace{14mu}{and}\mspace{14mu}{\psi_{2}\left( {\overset{\_}{x},\overset{\_}{y}} \right)}} = {\frac{\partial{\phi\left( {x,y} \right)}}{\partial y}.}}$The wavelet transform with respect to ψ₁ and ψ₂ is

$\begin{matrix}{{{T_{\psi}\lbrack f\rbrack}\left( {b,a} \right)} = \begin{pmatrix}{{T_{\psi_{1}}\lbrack f\rbrack} = {a^{- 2}{\int{d^{2}x\;{\psi_{1}\left( {a^{- 1}\left( {x - b} \right)} \right)}{f(x)}}}}} \\{{T_{\psi_{2}}\lbrack f\rbrack} = {a^{- 2}{\int{d^{2}x\;{\psi_{2}\left( {a^{- 1}\left( {x - b} \right)} \right)}{f(x)}}}}}\end{pmatrix}} \\{= {{{T_{\psi}\lbrack f\rbrack}\left( {b,a} \right)} = {\nabla\left\{ {{T_{\phi}\lbrack f\rbrack}\left( {b,a} \right)} \right\}}}}\end{matrix}$from which we can extract the modulus and argument of the WT:

T ψ ⁡ [ f ] ⁢ ( b , a ) = ( ψ ⁡ [ f ] ⁢ ( b , a ) , ψ ⁡ [ f ] ⁢ ( b , a ) ) ψ ⁡[ f ] ⁢ ( b , a ) = [ ( T ψ 1 ⁡ [ f ] ⁢ ( b , a ) ) 2 + ( T ψ 2 ⁡ [ f ] ⁢ ( b, a ) ) 2 ] 1 / 2 ψ ⁡ [ f ] ⁢ ( b , a ) = Arg ⁡ ( T ψ 1 ⁡ [ f ] ⁢ ( b , a ) +⁢T ψ 2 ⁡ [ f ] ⁢ ( b , a ) )

The wavelet transform modulus maxima are defined as the locations bwhere M_(ψ) [ƒ] (b,a) is locally maximum in the direction of theargument A_(ψ) [f] (b,a), at a given scale a>0. The WTMM lie onconnected chains and are thus called maxima chains (FIG. 7A-7F). One canthen find the maxima along these WTMM chains. The WTMM maxima, or WTMMMare defined as the points along the maxima chains where the M_(ψ) [ƒ](b,a) is locally maximum. The WTMMM are linked through scales to formthe space-scale skeleton (FIG. 7G). Hence, one can identify thesingularities of the function as the loci x where the WTMMM lines of theWT skeleton (FIG. 7G) point to in the limit a→0⁺. Along thesespace-scale vertical lines the WTMMM behave as a power-law˜a^(h(x))(M=ka^(h)) from which one can extract the local Hölder exponenth(x). The multifractal formalism amounts to characterize the relativecontributions of each Hölder exponent value via the estimate of theso-called D(h) singularity spectrum defined as the fractal dimension ofthe set of points x where h(x)=h. To compute D(h) we therefore usewavelets to partition the surface by defining the partition functiondirectly from the WTMMM in the skeleton:

⁢( q , a ) = ⁢ ( ⁢ ψ ⁡ [ f ] ⁢ ( x , a ′ ) ) qwhere L(a) is the set of all vertical space-scale lines in the skeletonthat exist at the given scale a>0 and which contain maxima at any scalea′≤a and q ε R. One can then define the exponent τ(q) from the power-lawbehavior of the partition function:

(q, a)˜a ^(τ(q)) , a→0⁺

-   And the D(h) singularity spectrum of ƒ can be determined from the    Legendre transform of the partition function scaling exponent    D(h)=min_(q)(qh−τ(q))-   In practice, to avoid instabilities in the estimation of the    singularity spectrum D(h) through the Legendre transform, we used h    and D(h) as mean quantities defined in a canonical ensemble, i.e.    with respect to their Boltzmann weights computed from the WTMMM:

W ψ ⁡ [ f ] ⁢ ( q , , a ) =  ⁢ ψ ⁡ [ f ] ⁢ ( x , a ⁢ ⁢ ′ )  q ⁢ ( q , a )Then one computes the expectation values:

h ⁡ ( q , a ) = ⁢ ln ⁢  ⁢ ψ ⁡ [ f ] ⁢ ( x , a ′ )  ⁢ W ψ ⁡ [ f ] ⁢ ( q , , a ). ⁢ D ⁡ ( q , a ) = ⁢ W ψ ⁡ [ f ] ⁢ ( q , , a ) ⁢ ln ⁡ ( W ψ ⁡ [ f ] ⁢ ( q , , a) )from which one derives

${h(q)} = {\frac{d\;{\tau(q)}}{dq} = {\lim_{a->0^{+}}{{{h\left( {q,a} \right)}/\ln}\; a}}}$D(q) = lim_(a− > 0⁺)D(q, a)/ln  aand thus the singularity spectrum D(h) as a curve parameterized by q.

Homogeneous monofractal functions with singularities of unique Hölderexponent H are characterized by a linear τ(q) curve of slope H. Anonlinear τ(q) is the signature of nonhomogeneous multifractalfunctions, meaning that the Hölder exponent is a fluctuating quantitythat depends on x. Then the corresponding singularity spectrum has acharacteristic single-humped shape. Note that for both mono- andmultifractal functionsD(q=0)=−τ(q=0)=D _(F)

-   Where D_(F) (noted simply D throughout the text) is the fractal    dimension of the support of singularities of f.    Methods—Statistical Tests

The Wilcoxon rank-sum test is a non-parametric statistical hypothesistest that is used as an alternative to Student's t-test when thepopulation cannot be assumed to be normally distributed. It was usedhere to calculate the p-values comparing the CC and MLO fractaldimensions and benign and malignant cases images since the benign datafollowed a bimodal distribution (FIG. 10). The calculations were doneusing the Wilcox test in R.

Methods—Bayesian Statistics

Bayes theorem states that

$\begin{matrix}{p\left( \theta \middle| G \right)} \\{posterior}\end{matrix} = {\begin{matrix}{p\left( \theta \middle| G \right)} \\{likelihood}\end{matrix}*{\begin{matrix}{p(\theta)} \\{prior}\end{matrix}/\begin{matrix}{p(G)} \\{evidence}\end{matrix}}}$where the prior, p(θ), represents the strength of our belief inmalignant lesions p(M) or benign lesions p(B) out of those that havebeen diagnosed by a radiologist. The posterior, p(θ|G), represents thestrength of our belief, having accounted for the geometrical evidence,G, where G represents the position of the lesion in the fractaldimension plot, either fractal (F) or Euclidean (E). The quotient of thelikelihood over the evidence, p(θ|G)/p(G), represents the support theevidence, G, provides for θ. Since the prior reflects uncertainty in theparameter value θ, p(θ) was based on a Beta distribution with aspecified mean and standard deviation. The Beta distribution is asfollows:

${f\left( {{\theta;\alpha},\beta} \right)} = {\frac{\Gamma\left( {\alpha + \beta} \right)}{{\Gamma(\alpha)}{\Gamma(\beta)}}{\theta^{\alpha - 1}\left( {1 - \theta} \right)}^{\beta - 1}}$

The Probability Model: To estimate the mean of the Beta distribution formalignant cases, p(M), the prevalence of mammograms with a BI-RADSassessment score of 3, 4, and 5 were used as determined by theradiologists diagnostics multiplied by the historical probability ofmammograms receiving those assessment scores result in malignant MCclusters respectively. One out of the 59 cases considered in this studyreceived an assessment score of 3, 47 out of 59 received a 4, and 11 outof 59 received a 5. Based on historical data, the probability ofmalignant lesions given an assessment score of 3 is 2%, an assessmentscore of 4 is 26.5% (taken as the midpoint of the reported interval of[23%-30%], and an assessment score of 5 is 95%. Therefore the Betadistribution for p(M) was chosen with a mean of

$\frac{{1*0.02} + {47*0.265} + {11*0.95}}{1 + 47 + 11} = 0.3885$and the Beta distribution for p(B) with a mean of 1-0.3885=0.6115.However, since there isn't much certainty regarding these values, theBeta distribution for p(M) and p(B) were assigned a relatively largestandard deviation of 0.25. This resulted in a Beta distribution forp(M) with parameters (α, β)=(1.09241,1.71223) and p(B) with parameters(α, β)=(1.71223,1.09241). The likelihood p(F|M), where F representsbreast lesions characterized as being in the fractal 465 zone (and Elikewise represents those in the Euclidean zone), is based on the 23 ofthe 25 malignant cases that were in the fractal zone; this likelihood isthe probability that the data could be generated with parameter valuesθ. Similarly, the likelihood p(E|B) is based on the 30 out of 34 benigncases that were in the Euclidean zone. To arrive at the posteriordistributions p(M|F) and p(B|E), the R routine “BernBeta.R” was used.

The resulting posterior distribution for p(MIF) was a Beta distributionwith parameters (α=24.0924, β=3.71223). Based on this distribution, theresulting 95% highest density interval was (0.742, 0.975. The posteriordistribution for P(B|E) was a Beta distribution with parameters(α=32.7122, β=5.09241). Based on this distribution, the resulting 95%highest density interval was (0.757, 0.962). The highest densityinterval spans 95% of the posterior distribution such that every pointinside the interval is deemed more credible. In other words, given theprior and the likelihood, observing the parameter value for thepercentage of breast lesions characterized in the fractal zone that aremalignant, there is a 95% probability that this parameter is between0.742 and 0.975. Similarly, for the percentage of breast lesionscharacterized in the Euclidean zone that are benign, there is a 95%probability that this parameter is between 0.757 and 0.962.

As described in detail in the 2D WTMM Method section and in FIG. 7, thewavelet transform (WT) acts as a mathematical microscope to characterizespatial image information over a continuous range of size scales. It isthe gradient vector of the image smoothed by dilated versions of aGaussian filter. At each size scale, the wavelet transform modulusmaxima (WTMM) are defined by the positions where the modulus of the WTis locally maximal. These WTMM are automatically organized as maximachains at the considered scale. Along each of these chains, furtherlocal maxima are found, i.e., the WTMM maxima (WTMMM). This process isrepeated for all size scales and the WTMMM from each scale are thenlinked to form the WT skeleton. As shown in FIGS. 8 and 9, the abilityto consider (vertical) space-scale WTMMM lines in the WT skeletonindividually is key, since it allows us to objectively discriminatebetween lines pointing to the tissue background from those pointing tothe microcalcifications by considering how the WT modulus varies as afunction of the scale parameter along each space-scale line. In FIGS. 8Dand 9D, each space-scale line obtained from the WT skeleton isrepresented by plotting the evolution of the WT modulus, M (see the 2DWTMM section above), as a function of the scale parameter, a, in alog-log plot. This relationship between M and a is characterized by apower-law behavior via the equationM=Ka^(h)where K is a pre-factor and h is the Holder exponent quantifying thestrength of the singularity to which the space-scale line is pointingto. In log-log plots shown in FIGS. 8D and 9D, the slope of the curvestherefore corresponds to h. By considering two types of informationcharacterizing the behavior of a space-scale line, namely the strengthof the modulus at the smallest scale, which is given by the log of thepre-factor, log(K), as well as the slope (in a logarithmicrepresentation) of the modulus variation across scales, h, aclassification procedure is setup which results in two sets ofspace-scale lines that clearly segregate MC from background tissue.Isolated MC can be seen as Dirac-like singularities through the opticsof the WT, for which h is theoretically known to be −1. However, whileclustered MC may not appear as isolated 200 Dirac-like singularities,the edge that they form is still easily detectable through thespace-scale lines, with a value of h˜0 (discontinuity). This means thatfor both isolated and clustered MC, we can expect the space-scale linesto behave as M =ka^(h) with h≤0, which contrasts from the healthybackground tissue, for which h˜1/3 for fatty breast tissue and h˜2/3 fordense breast tissue. However, relying only on h may not be sufficient,which is why the strength of the WT modulus at the lowest scale, whichquantifies the contrast between MC and background, is also needed. Forthe sample case presented in FIG. 8, the plot in FIG. 8D shows thatneither log(K) nor h, taken individually, would have been sufficient tosegregate between MC and background. However, for the sample casepresented in FIG. 9, the plot in FIG. 9D shows that log(K) alone wassufficient. A more detailed discussion of both cases follows.

In FIG. 8 the background breast tissue is dense, which makes thecontrast between background and MC weak (i.e. causing a low value forthe WT modulus of red curves at the smallest scale in FIG. 8D). However,the roughness fluctuations of dense breast tissue are characterized by arelatively high smoothness level, which translates to blue curves with alarge slope (i.e., a high h value, ˜2/3) for scales 10<a<200 pixels ascompared to the red curves with negative slopes for scales a≥10 pixelsthat correspond to WTMMM lines that point to MC at small scale (a→0+)(FIG. 8D). In FIG. 9, the background breast tissue is fatty, which ischaracterized by a higher roughness level (i.e. a lower h value ˜1/3,although still positive), that reduces the discriminatory power of h.However, for MC embedded into fatty tissue, the contrast is high, whichtranslates to a high value of log(K). Therefore, applying a threshold onboth parameters, h and log(K), is key to segregating MC from theirbackground regardless of the density (fatty or dense) of the compositionof the breast tissue.

Once this segregation is done, the so-called singularity spectrum canthen be calculated separately for each subset, which then allows us toconsider the fractal dimension D of the lesion, characterizing itsarchitecture.

We restricted the analysis of DDSM cases (see Methods sections above) toimages having a radiologist encircled region that was larger than 2562pixels for both views (CC and MLO) and also to make sure that thedistribution of patient ages was comparable (i.e. 56.7+/−11.4 years oldfor the benign cases and 65.5+/−12.4 years old for the malignant cases).This resulted in an analyzed sample with a total of 59 cases (118images), 34 of which are benign (68 images) and 25 of which aremalignant (50 images). The histograms of fractal dimension valuesobtained are presented in FIG. 10. Note that blending the CC and MLOfractal dimensions together in these distributions would not guaranteean unbiased statistical analysis, which is why the fractal dimensionvalues for the CC and MLO results were analyzed independently. FIG.10demonstrates that benign MC clusters have a strong preference forEuclidean dimensions that are either close to D=1 or to D=2 and thatthere is a very clear zone of avoidance in the fractal range, i.e., for1<D<2, with an actual gap in the benign histograms for the bin centeredat D=1.5 for both views. For the malignant cases, it is the opposite:Euclidean zones are avoided and the data are very clearly centered inthe fractal range for both views, with the peak of the histograms atD=1.5. A statistical comparison between benign vs. malignant MC clusterswas performed using the Wilcoxon rank-sum test, which yielded p-valuesof 0.009 for CC comparisons and 0.014 for MLO comparisons for the benignvs. malignant fractal dimension distributions. These are statisticallysignificant differences.

Example 6 The CC-MLO Fractal Dimension Plot and Fractal Zone

In this Example, a fractal dimension plot between the MLO and CC viewsof a breast is calculated. Unless otherwise specified, all samples,sample handling, and analysis was as described above in Example 5. Asshown in Example 5, the significance of the difference between benignand malignant is quite interesting. However, it is still only based onstatistics of populations. The histograms in FIG. 10 show that, wheneach view is taken independently (CC or MLO), it is still possible,though unlikely, for a malignant lesion to have a Euclidean dimension,and vice-versa, for benign lesions to have a (non-integer) fractaldimension. However, in order to work towards a potential CAD method thatwould be able to diagnose breast lesions individually instead of via thepopulation statistics, we combined the information to indirectly inferthe 3D structure of the tumor embedded into the breast tissue. This ispresented in a novel plot called the “CC-MLO fractal dimension plot”shown in FIG. 11, where red dots represent malignant cases and greendots represent benign cases. The square centered at (1.5, 1.5)represents those cases for which both CC and MLO views have a fractaldimension that is within 1.2<D<1.8. Note that only malignant cases arefound in this internal square. However, having one of the two views witha score that is close to D=1.5 may “compensate” for its other view beingoutside of the [1.2, 1.8]×[1.2, 1.8] square, i.e., as one viewapproaches D=1.5, the farther from 1.5 the other can be. Furtherjustification is presented below and in FIG. 12. That is how thetriangular regions that decay linearly as a function of distance fromthe internal square were defined. Therefore, the central square,combined with the four triangular regions extending from it are what wedefine as the “fractal zone”. Of the 59 cases considered in this study,92% of malignant breast lesions studied (23 out of 25) were in thefractal zone while 88% of the benign lesions were in the Euclidean zones(30 out of 34).

Bayesian Analysis

The inferences from a Bayesian analysis are richer and more informativethan null hypothesis significance testing. In particular, there is noreliance on p-values. But also, Bayesian models are designed to beappropriate to the data structure without having to make approximationassumptions typical in null hypothesis significance testing. The resultsreported above show that the vast majority of malignant breast lesionsare fractal, and that the vast majority of benign breast lesions areEuclidean. However, the condition of interest is how breast lesions inthe fractal zone can indicate malignancy, and how breast lesions in theEuclidean zone can indicate benignancy.

Bayesian inference derives the posterior probability as a consequence oftwo antecedents, a prior probability and a likelihood function derivedfrom a probability model for the data to be observed. In thisapplication, the model is based on historical radiology assessmentscores using the BI-RADS system. A detailed description of thisprobability model as well as the mathematical model behind Bayesanalysis is presented in the Methods section. Bayesian inference thenbecomes a computation of the posterior probability according to Bayes'rule. The interpretable output of this Bayesian analysis is theso-called 95% highest density interval (HDI), which is analogous to the95% confidence interval in frequentist statistics. The 95% HDI from theresulting posterior distribution indicates that the percentage of breastlesions in the fractal zone that are malignant is between 74.2% and97.5%. Alternatively, in terms of controlling for false positives, whichis a major concern, as discussed in the Introduction, the percentage ofbreast lesions in the Euclidean zone that are benign is between 75.7%and 96.2%.

Interpretation of the 3D Geometrical Structure

Even though two different 2D views of a 3D object are insufficient tofully characterize its 3D geometry, it can nonetheless give a robustestimate, as shown in this Example. The cases outside of the fractalzone can be categorized in two Euclidean subsets: 1) LINES, i.e. thosethat are approximately in the (D_(CC)=1,D_(MLO)=1) area, which aretherefore seen as one-dimensional objects from both views (FIG. 12A,left panel); or 2) SHEETS, i.e. those that are either in the (D_(CC)=1,D_(DMLO)=2) or (D_(CC)=2, D_(MLO)=1) areas, which are seen as a fulltwo-dimensional object in one view, but as a one-dimensional object fromthe other view and also those that are in the (D_(CC)=2, D_(MLO)=2)area, which are seen as a full two-dimensional object from both views(FIG. 12B, center panel). Although simplistic, these case modelsrepresent a good estimate of what the 3D Euclidean structure of a benignlesion may look-like.

For the cases that fall in the fractal zone, those malignant lesionsthat are in the [1.2,1.8]×[1.2,1.8] square have a fractal signature thatis seen from both views, whereas those that are in the triangular areaswould represent fractal clusters that grow onto a 2D plane, i.e. seen asa fractal from one view, but seen either as a line (bottom or lefttriangular regions) or a plane (top or right triangular regions) fromthe other view (FIG. 12C, right panel). Interestingly, adiffusion-limited aggregate embedded in 3D space and for which 2<D<3will have a 2D projection with D=2. Since no malignant lesions are foundin the (D_(CC)=2, D_(MLO)=2) area of the CC-MLO fractal dimension plot,we can safely hypothesize that all tumors are essentially limited to a2-dimensional fractal structure (within the 3-dimensional breasttissue), for which 1<D<2. Without wishing to be held to a particulartheory, this therefore leads us to conjecture that all breast tumorsconsidered in this study, benign and malignant, fractal or Euclidean,would grow on 2-dimensional manifolds.

Examples 5 and 6 illustrate embodiments of provided methodologies whichoffer a way to accurately classify benign and malignant tumors based ontheir invasiveness as determined by the geometrical structure. Byconsidering the organization of tumors via CAD systems, currentmammographic practice may be improved by increasing accuracy, andpotentially decreasing recall rates and costs. The inferred3-dimensional geometry of the breast lesions based on the analysis ofthe mammographic images using the 2D WTMM methodology allows one toexplore the invasiveness of the breast tumors and provide aninterpretation of the severity of the lesion. By considering where eachcase falls on the CC-MLO fractal dimension plot, a score similar to theBreast Imaging-Reporting and Data System (BI-RADS) assessment score maybe assigned to each case. Not only does this tool have the potential asa CAD, but it may also provide insight into the underlying mechanismsthat drive overall growth and structure at the time of the screeningmammogram.

Without wishing to be held to a particular theory, it is contemplatedthat, since the structure of the tumors are different, with benignlesions likely being smooth Euclidean objects and malignant lesionsbeing branching objects (and possibly, for both cases, being restrictedto growing along 2D manifolds within the 3D breast tissue environment),there may be a link to the cellular mechanisms at the lower levels inthe system that drive the organization at the much larger scale ofmammograms. Use of provided methods may be helpful in revealing theserelationships and improving detection and treatment.

EQUIVALENTS AND SCOPE

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. The scope of the presentinvention is not intended to be limited to the above Description, butrather is as set forth in the following claims:

We claim:
 1. A method comprising providing a first view of a region oftissue; providing a second view of the region of tissue; calculating afirst fractal dimension for the first view of the region of tissue; andcalculating a second fractal dimension for the second view of the regionof tissue; wherein a fractal zone is defined as a polygon consisting ofa central square and a first, second, third, and fourth extendingtriangular region as plotted on a graph of the fractal dimension of thefirst view of the region of tissue by the fractal dimension of thesecond view of the region of tissue; and wherein if the fractaldimension of at least one of the first fractal dimension and the secondfractal dimension is in the fractal zone, the region of tissue isconsidered cancerous.
 2. The method of claim 1, further comprisingtreating the region of tissue if it is cancerous.
 3. The method of claim1, wherein the central square and first, second, third and fourthextending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view: central square: (1.2,1.2), (1.2, 1.8), (1.8, 1.2)(1.8, 1.8); first extending triangularregion: (0.5, 1.5), (1.2, 1.2), (1.2, 1.8); second extending triangularregion: (1.5, 0.5), (1.2, 1.2), (1.8, 1.2); third extending triangularregion: (1.5, 2.3), (1.2, 1.8), (1.8, 1.8); and fourth extendingtriangular region: (2.3, 1.5), (1.8, 1.2), (1.8, 1.8).
 4. The method ofclaim 1, wherein the central square and first, second, third and fourthextending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view: central square: (1.1,1.1), (1.1, 1.9), (1.9, 1.1)(1.9, 1.9); first extending triangularregion: (0.5, 1.5), (1.1, 1.1), (1.1, 1.9); second extending triangularregion: (1.5, 0.5), (1.1, 1.1), (1.9, 1.1); third extending triangularregion: (1.5, 2.3), (1.1, 1.9), (1.9, 1.9); and fourth extendingtriangular region: (2.3, 1.5), (1.9, 1.1), (1.9, 1.9).
 5. The method ofclaim 1, wherein the central square and first, second, third and fourthextending triangular regions are defined by the following fractaldimensions, as plotted on a graph of the fractal dimension of the firstview by the fractal dimension of the second view: central square: (1.3,1.3), (1.3, 1.7), (1.7, 1.3)(1.7, 1.7); first extending triangularregion: (0.5, 1.5), (1.3, 1.3), (1.3, 1.7); second extending triangularregion: (1.5, 0.5), (1.3, 1.3), (1.7, 1.3); third extending triangularregion: (1.5, 2.3), (1.3, 1.7), (1.7, 1.7); and fourth extendingtriangular region: (2.3, 1.5), (1.7, 1.3), (1.7, 1.7).
 6. The method ofclaim 1, further comprising providing a third view of the region oftissue; and calculating a third fractal dimension for the third view ofthe region of tissue; wherein if the fractal dimension of at least oneof the first fractal dimension, the second fractal dimension, and thethird fractal dimension is in the fractal zone, the region of tissue isconsidered cancerous.
 7. The method of claim 1, wherein the tissue isselected from breast tissue, brain tissue, lung tissue, kidney tissue,liver tissue, uterine tissue, dermal tissue, and pancreatic tissue. 8.An apparatus comprising: a memory for storing a set of instructions; anda processor for executing the set of instructions, wherein theinstructions, when executed, cause the processor to: provide a firstview of a region of tissue; provide a second view of the region oftissue; calculate a first fractal dimension for the first view of theregion of tissue; and calculate a second fractal dimension for thesecond view of the region of tissue; wherein a fractal zone is definedas a polygon consisting of a central square and a first, second, third,and fourth extending triangular region as plotted on a graph of thefractal dimension of the first view of the region of tissue by thefractal dimension of the second view of the region of tissue; anddetermine if one or more of the fractal dimension and the second fractaldimension is in the fractal zone.
 9. The apparatus of claim 8, whereinthe central square and first, second, third and fourth extendingtriangular regions are defined by the following fractal dimensions, asplotted on a graph of the fractal dimension of the first view by thefractal dimension of the second view: central square: (1.2, 1.2), (1.2,1.8), (1.8, 1.2)(1.8, 1.8); first extending triangular region: (0.5,1.5), (1.2, 1.2), (1.2, 1.8); second extending triangular region: (1.5,0.5), (1.2, 1.2), (1.8, 1.2); third extending triangular region: (1.5,2.3), (1.2, 1.8), (1.8, 1.8); and fourth extending triangular region:(2.3, 1.5), (1.8, 1.2), (1.8, 1.8).
 10. The apparatus of claim 8,wherein the central square and first, second, third and fourth extendingtriangular regions are defined by the following fractal dimensions, asplotted on a graph of the fractal dimension of the first view by thefractal dimension of the second view: central square: (1.1, 1.1), (1.1,1.9), (1.9, 1.1)(1.9, 1.9); first extending triangular region: (0.5,1.5), (1.1, 1.1), (1.1, 1.9); second extending triangular region: (1.5,0.5), (1.1, 1.1), (1.9, 1.1); third extending triangular region: (1.5,2.3), (1.1, 1.9), (1.9, 1.9); and fourth extending triangular region:(2.3, 1.5), (1.9, 1.1), (1.9, 1.9).
 11. The apparatus of claim 8,wherein the central square and first, second, third and fourth extendingtriangular regions are defined by the following fractal dimensions, asplotted on a graph of the fractal dimension of the first view by thefractal dimension of the second view: central square: (1.3, 1.3), (1.3,1.7), (1.7, 1.3)(1.7, 1.7); first extending triangular region: (0.5,1.5), (1.3, 1.3), (1.3, 1.7); second extending triangular region: (1.5,0.5), (1.3, 1.3), (1.7, 1.3); third extending triangular region: (1.5,2.3), (1.3, 1.7), (1.7, 1.7); and fourth extending triangular region:(2.3, 1.5), (1.7, 1.3), (1.7, 1.7).
 12. The apparatus of claim 8,wherein the processor further: provides a third view of the region oftissue; and calculates a third fractal dimension for the third view ofthe region of tissue.
 13. The apparatus of claim 8, wherein the tissueis selected from breast tissue, brain tissue, lung tissue, kidneytissue, liver tissue, uterine tissue, dermal tissue, and pancreatictissue.